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In flies (Diptera), the ovary displays several distinct patterns of the follicular epithelium formation and diversification. Two main patterns have been identified in the true flies or Brachycera, namely the Rhagio type and the Drosophila type. These patterns align with the traditional division of Brachycera into Orthorrhapha and Cyclorrhapha. However, studies of the follicular epithelium morphogenesis in cyclorrhaphans other than Drosophila are scarce. We characterise the developmental changes associated with the emergence of follicle cell (FC) diversity in two cyclorrhaphans belonging to the family Tephritidae (Brachycera, Cyclorrhapha). Our analysis revealed that the diversification of FCs in these species shows characteristics of both the Rhagio and Drosophila types. First, a distinct cluster of FCs, consisting of polar cells and border‐like cells, differentiates at the posterior pole of the ovarian follicle. This feature is unique to the Rhagio type and has only been reported in species representing the Orthorrhapha group. Second, morphological criteria have identified a significantly smaller number of subpopulations of FCs than in Drosophila. Furthermore, while the general pattern of FC migration is similar to that of Drosophila, the distinctive migration of the anterior‐dorsal FCs is absent. In the studied tephritids, the migration of the anterior polar cell/border cell cluster towards the anterior pole of the oocyte is followed by the posterior migration of the main body cuboidal FCs to cover the expanding oocyte. Finally, during the onset of vitellogenesis, a distinct subset of FCs migrates towards the centre of the ovarian follicle to cover the oocyte's anterior pole. Our study also highlights specific actions of some FCs that accompany the migration process, which has not been previously documented in cyclorrhaphans. These results support the hypothesis that the posterior and centripetal migrations of morphologically unique FC subsets arose in the common ancestor of Cyclorrhapha. These events appear to have occurred fairly recently in the evolutionary timeline of Diptera. [ABSTRACT FROM AUTHOR]

The Drosophila egg chamber (EC) starts as a spherical tissue at the beginning. With maturation, the outer follicle cells of EC collectively migrate in a direction perpendicular to the anterior-posterior axis, to shape EC from spherical to ellipsoidal. Filamentous actin (F-actin) plays a significant role in shaping individual migratory cells to the overall EC shape, like in every cell migration. The primary focus of this article is to unveil the function of different Actin Binding Proteins (ABPs) in regulating mature Drosophila egg shape. We have screened 66 ABPs, and the genetic screening data revealed that individual knockdown of Arp2/3 complex genes and the “capping protein β” (cpb) gene have severely altered the egg phenotype. Arpc1 and cpb RNAi mediated knockdown resulted in the formation of spherical eggs which are devoid of dorsal appendages. Studies also showed the role of Arpc1 and cpb on the number of laid eggs and follicle cell morphology. Furthermore, the depletion of Arpc1 and cpb resulted in a change in F-actin quantity. Together, the data indicate that Arpc1 and cpb regulate Drosophila egg shape, F-actin management, egg-laying characteristics and dorsal appendages formation. [ABSTRACT FROM AUTHOR]

In nearly every species of insect, embryonic development takes place outside of the mother's body and is entirely dependent on the elements that the mother had previously stored within the eggs. It is well known that the follicle cells (FCs) synthesize the eggshell (chorion) components during the process of choriogenesis, the final step of oogenesis before fertilization. These cells have developed a specialization in the massive production of chorion proteins, which are essential for the protection and survival of the embryo. Here, we investigate the function of Sec16, a protein crucial for the endoplasmic reticulum (ER) to Golgi traffic, in the oocyte development in the insect Rhodnius prolixus. We discovered that Sec16 is strongly expressed in vitellogenic females' ovaries, particularly in the choriogenic oocyte and it is mainly associated with the FCs. Silencing of Sec16 by RNAi caused a sharp decline in oviposition rates, F1 viability, and longevity in adult females. In the FCs, genes involved in the unfolded protein response (UPR), the ubiquitin-proteasome system (UPS), and autophagy were massively upregulated, whereas the mRNAs of Rp30 and Rp45-which code for the two major chorion proteins - were downregulated as a result of Sec16 silencing, indicating general proteostasis disturbance. As a result, the outer surface ultrastructure of Sec16-silenced chorions was altered, with decreased thickness, dityrosine crosslinking, sulfur signals, and lower amounts of the chorion protein Rp30. These findings collectively demonstrate the critical role Sec16 plays in the proper functioning of the FCs, which impacts the synthesis and deposition of particular components of the chorion as well as the overall reproduction of this vector. [ABSTRACT FROM AUTHOR]

In nearly every species of insect, embryonic development takes place outside of the mother’s body and is entirely dependent on the elements that the mother had previously stored within the eggs. It is well known that the follicle cells (FCs) synthesize the eggshell (chorion) components during the process of choriogenesis, the final step of oogenesis before fertilization. These cells have developed a specialization in the massive production of chorion proteins, which are essential for the protection and survival of the embryo. Here, we investigate the function of Sec16, a protein crucial for the endoplasmic reticulum (ER) to Golgi traffic, in the oocyte development in the insect Rhodnius prolixus. We discovered that Sec16 is strongly expressed in vitellogenic females’ ovaries, particularly in the choriogenic oocyte and it is mainly associated with the FCs. Silencing of Sec16 by RNAi caused a sharp decline in oviposition rates, F1 viability, and longevity in adult females. In the FCs, genes involved in the unfolded protein response (UPR), the ubiquitin-proteasome system (UPS), and autophagy were massively upregulated, whereas the mRNAs of Rp30 and Rp45—which code for the two major chorion proteins - were downregulated as a result of Sec16 silencing, indicating general proteostasis disturbance. As a result, the outer surface ultrastructure of Sec16-silenced chorions was altered, with decreased thickness, dityrosine crosslinking, sulfur signals, and lower amounts of the chorion protein Rp30. These findings collectively demonstrate the critical role Sec16 plays in the proper functioning of the FCs, which impacts the synthesis and deposition of particular components of the chorion as well as the overall reproduction of this vector.

The follicle stem cells (FSCs) in the Drosophila ovary are an important experimental model for the study of epithelial stem cell biology. Although decades of research support the conclusion that there are two FSCs per ovariole, a recent study used a novel clonal marking system to conclude that there are 15-16 FSCs per ovariole. We performed clonal analysis using both this novel clonal marking system and standard clonal marking systems, and identified several problems that may have contributed to the overestimate of FSC number. In addition, we developed new methods for accurately measuring clone size, and found that FSC clones produce, on average, half of the follicle cells in each ovariole. Our findings provide strong independent support for the conclusion that there are typically two active FSCs per ovariole, though they are consistent with up to four FSCs per germarium.

Epithelial cells form continuous membranous structures for organ formation, and these cells are classified into three major morphological categories: cuboidal, columnar, and squamous. It is crucial that cells transition between these shapes during the morphogenetic events of organogenesis, yet this process remains poorly understood. All three epithelial cell shapes can be found in the follicular epithelium of Drosophila egg chamber during oogenesis. Squamous cells (SCs) are initially restricted to the anterior terminus in cuboidal shape. They then rapidly become flattened to assume squamous shape by stretching and expansion in 12 ​h during midoogenesis. Previously, we reported that Notch signaling activated a zinc-finger transcription factor Broad (Br) at the end of early oogenesis. Here we report that ecdysone and JAK/STAT pathways subsequently converge on Br to serve as an important spatiotemporal regulator of this dramatic morphological change of SCs. The early uniform pattern of Br in the follicular epithelium is directly established by Notch signaling at stage 5 of oogenesis. Later, ecdysone and JAK/STAT signaling activities synergize to suppress Br in SCs from stage 8 to 10a, contributing to proper SC squamous shape. During this process, ecdysone signaling is essential for SC stretching, while JAK/STAT regulates SC clustering and cell fate determination. This study reveals an inhibitory role of ecdysone signaling in suppressing Br in epithelial cell remodeling. In this study we also used single-cell RNA sequencing data to highlight the shift in gene expression which occurs as Br is suppressed and cells become flattened. [Display omitted] • Notch, ecdysone and JAK/STAT signaling controls Broad to change the epithelial shape. • Ecdysone signaling regulates follicle cell stretching and dispersion. • JAK/STAT ensures proper spacing between cells to prevent clustering. • Single-cell RNA sequencing data reveal genes expressed during cell shape change. [ABSTRACT FROM AUTHOR]

Drosophila oogenesis requires the subsequent growth of distinct egg chambers each containing a group of sixteen germline cells surrounded by a simple epithelium of follicle cells. The oocyte occupies a posterior position within the germ cells, thus giving a distinct asymmetry to the egg chamber. Although this disposition is critical for the formation of the anterior–posterior axis of the embryo, the interplay between somatic and germ cells during the early stages of oogenesis remains an open question. We uncover by stage 2, when the egg chambers leaved the germarium, some unique spatial interactions between the posterior follicle cells and the oocyte. These interactions are restricted to the surface of the oocyte over the centriole cluster that formed during early oogenesis. Moreover, the posterior follicle cells in front of the oocyte display a convoluted apical membrane with extensive contacts, whereas the other follicle cells have a flat apical surface without obvious surface protrusions. In addition, the germ cells located at the posterior end of the egg chamber have very elongated protrusions that come into contact with each other or with facing follicle cells. These observations point to distinct polarization events during early oogenesis supporting previous molecular data of an inherent asymmetry between the anterior and the posterior regions of the egg chambers. [ABSTRACT FROM AUTHOR]

Apicobasal cell polarity loss is a founding event in epithelial–mesenchymal transition and epithelial tumorigenesis, yet how pathological polarity loss links to plasticity remains largely unknown. To understand the mechanisms and mediators regulating plasticity upon polarity loss, we performed single-cell RNA sequencing of Drosophila ovaries, where inducing polarity-gene l(2)gl-knockdown (Lgl-KD) causes invasive multilayering of the follicular epithelia. Analyzing the integrated Lgl-KD and wildtype transcriptomes, we discovered the cells specific to the various discernible phenotypes and characterized the underlying gene expression. A genetic requirement of Keap1-Nrf2 signaling in promoting multilayer formation of Lgl-KD cells was further identified. Ectopic expression of Keap1 increased the volume of delaminated follicle cells that showed enhanced invasive behavior with significant changes to the cytoskeleton. Overall, our findings describe the comprehensive transcriptome of cells within the follicle cell tumor model at the single-cell resolution and identify a previously unappreciated link between Keap1-Nrf2 signaling and cell plasticity at early tumorigenesis.

BackgroundGerm cells are specified during early development and are responsible for generating gametes in the adult. After germ cells are specified, they typically migrate to a particular niche in the organism where they reside for the remainder of its lifetime. For some model organisms, the specification and migration of germ cells have been extensively studied, but how these events occur in animals that reproduce both sexually and asexually is not well understood.ResultsWe have identified a novel TGF-β family member in Botryllus schlosseri, tgfβ-f, and found that it is expressed by follicle cell progenitors and the differentiated follicle and support cells surrounding the maturing gametes. Using the expression of tgfβ-f and the germ cell marker vasa, we have found that nearly all germ cells in Botryllus are associated with tgfβ-f-expressing follicle progenitors in clusters consisting solely of those two cell types. These clusters were mostly small, consisting of ten or fewer cells, and generally contained between a 2:1 and 1:1 ratio of follicle progenitors to germ cells. Clusters of germ and follicle progenitor cells were primarily localized to niches in the primary and secondary buds, but could also be found in other locations including the vasculature. We analyzed the location of germ cell clusters throughout the asexual life cycle of Botryllus and found that at the stage when germ cells are first detected in the secondary bud niche, a dramatic change in the size and location of germ/follicle cell clusters also occurred.ConclusionsOur findings suggest that germ/follicle cell clusters have predictable migratory patterns during the weekly asexual developmental cycle in Botryllus. An increased number of small clusters and the presence of clusters in the vasculature coinciding with the appearance of clusters in the secondary bud suggest that fragmentation of clusters and the migration of smaller clusters through the vasculature may be an important aspect of Botryllus reproductive biology, ensuring the transmission of the germline to subsequent asexual generations.

Cell polarity is a pre-requirement for many fundamental processes in animal cells, such as asymmetric cell division, axon specification, morphogenesis and epithelial tissue formation. For all these different processes, polarization is established by the same set of proteins, called partitioning defective (Par) proteins. During development in Drosophila melanogaster, decision making on the cellular and organism level is achieved with temporally controlled cell polarization events. The initial polarization of Par proteins occurs as early as in the germline cyst, when one of the 16 cells becomes the oocyte. Another marked event occurs when the anterior–posterior axis of the future organism is defined by Par redistribution in the oocyte, requiring external signaling from somatic cells. Here, we review the current literature on cell polarity events that constitute the oogenesis from the stem cell to the mature egg.

In insects, the follicle cells (FCs) give rise to a single-layered tissue of binucleated professional secretory cells that surround the oocytes during oogenesis. In the latest stage of oocyte development, the FCs rapidly synthesize and secrete the chorion (eggshell) immediately before degenerating through apoptosis. Here, we used RT-qPCR, electron microscopy, and RNAi silencing to explore the role of the main unfolded protein response (UPR) receptors IRE1 and PERK, as well as the ultrastructure dynamics of the FCs during oogenesis of the insect vector of Chagas disease Rhodnius prolixus. We found that IRE1 and PERK mRNAs are highly expressed in the ovaries of vitellogenic females. Interestingly, we observed that IRE1 and PERK, as well as different isoforms of the chaperones Bip and PDI, have their FCs gene expression levels decreased during the vitellogenesis to choriogenesis transition. Using transmission electron microscopy, we observed that the downregulation of the UPR gene expression is accompanied by dramatic changes in the FCs ultrastructure, with an 80% reduction in the mean area of the ER tubules, and circularization and enlargement of the mitochondria. Additionally, we found that parental RNAi silencing of both IRE1 and PERK resulted in minor changes in the chorion protein composition and ultrastructure, accessed by urea extraction of the chorion proteins and scanning electron microscopy, respectively, but did not impact the overall levels of oviposition and F1 embryo development. [ABSTRACT FROM AUTHOR]

Oogenesis is a fundamental biological process observed in most animals. Animals employ many different mechanisms to make their eggs and oogenesis cannot be completed without the support of other intraovarian and/or extraovarian cells and tissues. Invertebrates account for 99% of all animal life on earth and their complex life histories are a reflection of the structural heterogeneity of their ovaries and the diversity of their vitellogenic mechanisms. The process of oogenesis in different invertebrates differs because it is constrained by limits imposed by their respective body design, the architecture of their ovaries and their collective phylogenetic history. Oogenesis in the majority of invertebrates has yet to be investigated and current knowledge is limited to relatively superficial descriptions of a fraction of 1% of described species in each of the major animal phyla. It has been over 30 years since the last published review of invertebrate oogenesis and during the intervening years considerable progress has been made in our understanding of the ultrastructural, biochemical and molecular aspects of the process. This review aims to identify some unifying themes by collating and synthesizing the widely dispersed literature on invertebrate oogenesis covering 30 metazoan phyla for which basic information is available. [ABSTRACT FROM AUTHOR]

Gamete production in mammals and insects is controlled by cell signaling pathways that facilitate communication between germ cells and somatic cells. Nuclear receptor signaling is a key mediator of many aspects of reproduction, including gametogenesis. For example, the NR5A subfamily of nuclear receptors is essential for gonad development and sex steroid production in mammals. Despite the original identification of the NR5A subfamily in the model insect Drosophila melanogaster, it has been unclear whether Drosophila NR5A receptors directly control oocyte production. Ftz-f1 is expressed throughout the ovary, including in germline stem cells, germline cysts, and several populations of somatic cells. We show that ftz-f1 is required in follicle cells prior to stage 10 to promote egg chamber survival at the midoogenesis checkpoint. Our data suggest that egg chamber death in the absence of ftz-f1 is due, at least in part, to failure of follicle cells to exit the mitotic cell cycle or failure to accumulate oocyte-specific factors in the germline. Taken together, these results show that, as in mammals, the NR5A subfamily promotes maximal reproductive output in Drosophila. Our data underscore the importance of nuclear receptors in the control of reproduction and highlight the utility of Drosophila oogenesis as a key model for unraveling the complexity of nuclear receptor signaling in gametogenesis [ABSTRACT FROM AUTHOR]

In insects, synthesis and deposition of the chorion (eggshell) are performed by the professional secretory follicle cells (FCs) that surround the oocytes in the course of oogenesis. Here, we found that ULK1/ATG1, an autophagy‐related protein, is highly expressed in the FCs of the Chagas‐Disease vector Rhodnius prolixus, and that parental RNAi silencing of ULK1/ATG1 results in oocytes with abnormal chorion ultrastructure and FCs presenting expanded rough ER membranes as well as increased expression of the ER chaperone BiP3, both indicatives of ER stress. Silencing of LC3/ATG8, another essential autophagy protein, did not replicate the ULK1/ATG1 phenotypes, whereas silencing of SEC16A, a known partner of the noncanonical ULK1/ATG1 function in the ER exit sites phenocopied the silencing of ULK1/ATG1. Our findings point to a cooperated function of ULK1/ATG1 and SEC16A in the FCs to complete choriogenesis and provide additional in vivo phenotype‐based evidence to the literature of the role of ULK1/ATG1 in the ER in a professional secretory cell. [ABSTRACT FROM AUTHOR]

The follicle stem cells (FSCs) in the Drosophila ovary are an important experimental model for the study of epithelial stem cell biology. Although decades of research support the conclusion that there are two FSCs per ovariole, a recent study used a novel clonal marking system to conclude that there are 15–16 FSCs per ovariole. We performed clonal analysis using both this novel clonal marking system and standard clonal marking systems, and identified several problems that may have contributed to the overestimate of FSC number. In addition, we developed new methods for accurately measuring clone size, and found that FSC clones produce, on average, half of the follicle cells in each ovariole. Our findings provide strong independent support for the conclusion that there are typically two active FSCs per ovariole, though they are consistent with up to four FSCs per germarium.

The aim of this paper is to assess, by real‐time polymerase chain reaction and in situ hybridization, the expression of estrogen receptors ER1 and ER2 during the ovarian cycle of Mytilus galloprovincialis. By considering four phases of the reproductive cycle, that is stasis and previtellogenic stage (Stage 0), early vitellogenesis (Stage I), vitellogenesis (Stage II), full‐grown oocyte (Stage III), our investigation demonstrates that the two receptors are differently expressed during the phases investigated of the ovarian cycle: ER1 reaches the highest level at Stage III, whereas ER2 reaches the highest level at Stage II, with ER2 always present at higher levels than ER1. The stage‐dependent receptor expression was recorded within oocytes, follicle cells, and adipogranular cells. No ER1 and ER2 messenger RNAs (mRNAs) were found within vesicular cells. It is to be noted that the ER1 and ER2 expression within the growing oocytes, the follicular, and adipogranular cells overlaps with that of the mRNA for vitellogenin in the same cells, strongly suggesting that in Mytilus, as in vertebrates studied so far, the vitellogenin expression is under the control of estrogens. Research Highlights: The aim of this paper is to assess, by real‐time polymerase chain reaction and in situ hybridization, the expression of estrogen receptors ER1 and ER2 during the ovarian cycle of Mytilus galloprovincialisWe demonstrated that the two receptors are differently expressed during the phases investigated of the ovarian cycle: ER1 reaches the highest level at Stage III (full‐grown oocyte), whereas ER2 reaches the highest level at Stage II (vitellogenesis), with ER2 always present at higher levels than ER1. [ABSTRACT FROM AUTHOR]

Multiple aspects of Drosophila oogenesis, including germline stem cell activity, germ cell differentiation, and follicle survival, are regulated by the steroid hormone ecdysone. While the transcriptional targets of ecdysone signaling during development have been studied extensively, targets in the ovary remain largely unknown. Early studies of salivary gland polytene chromosomes led to a model in which ecdysone stimulates a hierarchical transcriptional cascade, wherein a core group of ecdysone-sensitive transcription factors induce tissue-specific responses by activating secondary branches of transcriptional targets. More recently, genome-wide approaches have identified hundreds of putative ecdysone-responsive targets. Determining whether these putative targets represent bona fide targets in vivo, however, requires that they be tested via traditional mutant analysis in a cell-type specific fashion. To investigate the molecular mechanisms whereby ecdysone signaling regulates oogenesis, we used genetic mosaic analysis to screen putative ecdysone-responsive genes for novel roles in the control of the earliest steps of oogenesis. We identified a cohort of genes required for stem cell maintenance, stem and progenitor cell proliferation, and follicle encapsulation, growth, and survival. These genes encode transcription factors, chromatin modulators, and factors required for RNA transport, stability, and ribosome biogenesis, suggesting that ecdysone might control a wide range of molecular processes during oogenesis. Our results suggest that, although ecdysone target genes are known to have cell type-specific roles, many ecdysone response genes that control larval or pupal cell types at developmental transitions are used reiteratively in the adult ovary. These results provide novel insights into the molecular mechanisms by which ecdysone signaling controls oogenesis, laying new ground for future studies.

Abstract Background Germ cells are specified during early development and are responsible for generating gametes in the adult. After germ cells are specified, they typically migrate to a particular niche in the organism where they reside for the remainder of its lifetime. For some model organisms, the specification and migration of germ cells have been extensively studied, but how these events occur in animals that reproduce both sexually and asexually is not well understood. Results We have identified a novel TGF-β family member in Botryllus schlosseri, tgfβ-f, and found that it is expressed by follicle cell progenitors and the differentiated follicle and support cells surrounding the maturing gametes. Using the expression of tgfβ-f and the germ cell marker vasa, we have found that nearly all germ cells in Botryllus are associated with tgfβ-f-expressing follicle progenitors in clusters consisting solely of those two cell types. These clusters were mostly small, consisting of ten or fewer cells, and generally contained between a 2:1 and 1:1 ratio of follicle progenitors to germ cells. Clusters of germ and follicle progenitor cells were primarily localized to niches in the primary and secondary buds, but could also be found in other locations including the vasculature. We analyzed the location of germ cell clusters throughout the asexual life cycle of Botryllus and found that at the stage when germ cells are first detected in the secondary bud niche, a dramatic change in the size and location of germ/follicle cell clusters also occurred. Conclusions Our findings suggest that germ/follicle cell clusters have predictable migratory patterns during the weekly asexual developmental cycle in Botryllus. An increased number of small clusters and the presence of clusters in the vasculature coinciding with the appearance of clusters in the secondary bud suggest that fragmentation of clusters and the migration of smaller clusters through the vasculature may be an important aspect of Botryllus reproductive biology, ensuring the transmission of the germline to subsequent asexual generations.

LIM domain, constituted by two tandem C2H2 zinc finger motif, proteins regulate several biological processes. They are usually found associated with various functional domains like Homeodomain, kinase domain and other protein binding domains. LIM proteins that are devoid of other domains are called LIM only proteins (LMO). LMO proteins were first identified in humans and are implicated in development and oncogenesis. They regulate various cell specifications by regulating the activity of respective transcriptional complexes. The Drosophila LMO protein (dLMO), Beadex (Bx), regulates various developmental processes like wing margin development and bristle development. It also regulates Drosophila behavior in response to cocaine and ethanol. We have previously generated Bx null flies and shown its essential function in neurons for multiple aspects of female reproduction. However, it was not known whether Bx affects reproduction through its independent function in ovaries. In this paper we show that female flies null for Bx lay eggs with multiple defects. Further, through knock down studies we demonstrate that function of Bx in follicle cells is required for normal egg development. We also show that function of Bx is particularly required in border cells for Drosophila fertility. [ABSTRACT FROM AUTHOR]

Deciphering the evolution of morphological structures is a remaining challenge in the field of developmental biology. The respiratory structures of insect eggshells, called the dorsal appendages, provide an outstanding system for exploring these processes since considerable information is known about their patterning and morphogenesis in Drosophila melanogaster and dorsal appendage number and morphology vary widely across Drosophilid species. We investigated the patterning differences that might facilitate morphogenetic differences between D. melanogaster, which produces two oar-like structures first by wrapping and then elongating the tubes via cell intercalation and cell crawling, and Scaptodrosophila lebanonensis, which produces a variable number of appendages simply by cell intercalation and crawling. Analyses of BMP pathway components thickveins and P-Mad demonstrate that anterior patterning is conserved between these species. In contrast, EGF signaling exhibits significant differences. Transcripts for the ligand encoded by gurken localize similarly in the two species, but this morphogen creates a single dorsolateral primordium in S. lebanonensis as defined by activated MAP kinase and the downstream marker broad. Expression patterns of pointed, argos, and Capicua, early steps in the EGF pathway, exhibit a heterochronic shift in S. lebanonensis relative to those seen in D. melanogaster. We demonstrate that the S. lebanonensis Gurken homolog is active in D. melanogaster but is insufficient to alter downstream patterning responses, indicating that Gurken-EGF receptor interactions do not distinguish the two species’ patterning. Altogether, these results differentiate EGF signaling patterns between species and shed light on how changes to the regulation of patterning genes may contribute to different tube-forming mechanisms. [ABSTRACT FROM AUTHOR]

In insects that lay eggs in large clutches, yolk accumulation in each of the many ovarioles is restricted to the basal (terminal) oocyte, the one closest to the lateral oviduct. All succeeding (subterminal) oocytes remain small until the terminal oocytes finished their development and were ovulated into the oviduct. The major step regulating yolk uptake by terminal oocytes is the formation of gaps between cells of the follicle layer, a process termed patency. In the migratory as well as in the desert locust, patency is induced by a Patency Inducing Factor (PIF) produced by the lateral oviducts. PIF is secreted in all regions of the lateral oviducts and interacts with the basal follicle cells via the pedicel, a fine duct that connects an ovariole with the oviduct. By this mechanism, patency is triggered in the follicle cells of the terminal oocyte only, restricting yolk accumulation to the oocytes next to ovulation. In contrast to the previous hypothesis, juvenile hormone (JH) is not necessary to induce patency, rather JH amplifies the effect of PIF. [ABSTRACT FROM AUTHOR]

Somatic follicle cells are critical support cells for Drosophila oogenesis, as they provide signals and molecules needed to produce a mature egg. Throughout this process, the follicle cells differentiate into multiple subpopulations and transition between three different cell cycle programs to support nurse cell and oocyte development. The follicle cells are mitotic in early egg chamber development, as they cover the germline cyst. In mid-oogenesis, follicle cells switch from mitosis to endocycling, increasing their ploidy from 2C to 16C. Finally, in late oogenesis, cells transition from endocycling to gene amplification, increasing the copy number of a small subset of genes, including the genes encoding proteins required for egg maturation. In order to explore the genetic regulation of these cell cycle switches and follicle cell development and specification, clonal analysis and the GAL4/UAS system are used frequently to reduce or increase expression of genes of interest. These genetic approaches combined with immunohistochemistry and in situ hybridization are powerful tools for characterizing the mechanisms regulating follicle cell development and the mitosis/endocycle and endocycle/gene amplification transitions. This chapter describes the genetic tools available to manipulate gene expression in follicle cells, as well as the methods and reagents that can be utilized to explore gene expression throughout follicle cell development., (© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Reproductive division in termites is the most significant biological process that leads to the formation of caste-specific differences in tasks and status. However, little is known about the mechanism of reproductive division that underlies caste differentiation. In the present study, ovarian development and stage-specific apoptotic patterns are investigated during oogenesis in reproductive, worker and soldier termites Reticulitermes aculabialis Tsai & Hwang. The results show that the mean lengths of the ovaries of reproductives are two-fold longer compared with those of workers and six-fold longer compared with soldiers. By contrast to the reproductives, the process of oogenesis in the workers includes only the oogonium differentiation stage (stage I) and oocyte growth stage (stage II), and oogenesis in the soldiers stops at stage I. Vitelogenic oocytes (stage III) are absent from workers and soldiers. During stage II in the reproductives and workers, the layer of follicle cells has a thickness of 7.56 ± 0.52 and 2.81 ± 0.34 µm, respectively. In addition, there are significant differences in the number and size of the germ cells at the same stage in the various castes. The existence of two apoptotic patterns during oogenesis is demonstrated by the terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling ( TUNEL) assay. First, the majority of the cells showing apoptosis occur at stage I of oogenesis in reproductives, workers and soldiers. Second, DNA fragmentation is demonstrated by TUNEL staining of the follicle cell layers and oocytes at stage II in reproductives. Finally, the proliferation activity of follicle cells in the reproductives is observed by 5-bromo-2′-deoxy-uridine labelling. The level of oogenesis may explain the significant discrepancies in the reproductive capacity among the reproductives, soldiers and workers. These large discrepancies are controlled by apoptosis during early oogenesis. [ABSTRACT FROM AUTHOR]

In addition to the reported single c DNA sequence encoding vitellogenin in the Pacific oyster Crassostrea gigas (referred as Cg-Vtg1), we isolated and characterized the 3′ end of a novel vitellogenin (Vtg) in C. gigas, named Cg-Vtg2. The transcript size is up to 6 kb long and includes a D-type von Willebrand domain, characteristic of most vitellogenins, but not occurring in Cg-Vtg1. The expression of Cg-Vtg2 was found restricted to follicular cells in maturing and ripe female gonads. Quantitative expression analysis of both vitellogenins showed a closely correlated transcription pattern during the female gametogenic cycle, with the highest expression during the ripe (mature) stage, and a declining expression, to almost undetectable levels, in post-spawning (resting) stage. Levels of vitellogenin m RNA associates with the quantity of ingested food. Mature female oysters fed either a 2% or a 12% algal biomass per female biomass, showed a 2.9-fold increase in expression of Cg-Vtg1 and a 3.7-fold increase of Cg-Vtg2 expression levels in those females fed the higher level of algae, suggesting that Vtg's could be used as quantitative indicator of vitellogenesis. Our results suggest that vitellogenins are synthesized during the late period of vitellogenesis, when most of the yolk has already been incorporated into maturing oocytes. [ABSTRACT FROM AUTHOR]

Abstract: The adverse effects of endocrine-disrupting chemicals (EDCs) have been well documented; however, the action mechanisms of many EDCs remain elusive and controversial. Furthermore, the highly diversified chemical structures and low environmental concentrations of EDCs present a major challenge to their chemical detection. Clearly, there is an urgent need for simple and reliable bioassays to detect EDCs in the environment and unravel their action mechanisms. We have recently identified luteinizing hormone receptor (lhcgr) as a robust estradiol (E2)-responsive gene in cultured zebrafish ovarian follicle cells. The expression of lhcgr exhibited a distinct biphasic response to E2 over a 24-h time-course treatment, making this a unique system for characterizing estrogenic EDCs. This study was undertaken to validate this platform by testing a wide range of EDCs, including 17α-ethinylestradiol (EE2), diethylstilbestrol (DES), bisphenol A (BPA), genistein (GEN), 1,1,1-trichloro-2-(2-chlorophenyl)-2-(4-chlorophenyl)ethane (o,p′-DDT), vinclozolin (VIN), bis(2-ethylhexyl) phthalate (DEHP), 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), and 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47). Diethylstilbestrol (DES), EE2 and o,p′-DDT mimicked E2 and induced a biphasic expression of lhcgr while BPA and GEN stimulated a monophasic expression in the 24-h time-course. In contrast, BDE-47, DEHP and VIN had no effect, whereas TCDD decreased lhcgr expression. Dose–response experiment showed that E2, EE2 and DES had the highest potency, which was followed by GEN, BPA and o,p′-DDT. The effects of estrogenic EDCs were further confirmed by their potentiation of hCG-induced activin βA2 subunit (inhbab) expression. In conclusion, the present study showed that the expression of lhcgr in cultured zebrafish follicle cells and its biphasic response to estrogens provide a unique in vitro platform for screening and categorizing estrogenic substances and deciphering their action mechanisms. [Copyright &y& Elsevier]

The study was aimed at understanding the process of reproduction and the changes happening in the ovary of Portunus pelagicus during maturation, which would be useful for its broodstock development for hatchery purposes. For that, tissue samples from different regions of the ovary at various stages of maturation were subjected to light and electron microscopy, and based on the changes revealed and the differences in ovarian morphology, the ovary was divided into five stages such as immature (previtellogenic oocytes), early maturing (early vitellogenic oocytes), late maturing (late vitellogenic oocytes), mature (vitellogenic oocytes), and spent (resorbing oocytes). The ovarian wall comprised of an outermost thin pavement epithelium, a middle layer of connective tissue, and an innermost layer of germinal epithelium. The oocytes matured as they moved from the centrally placed germinal zone toward the ovarian wall. The peripheral arrangement of nucleolar materials and the high incidence of cell organelles during the initial stages indicated vitellogenesis I. Movement of follicle cells toward oocytes in the early maturing stage and low incidence of mitochondria and endoplasmic reticulum in the ooplasm during late vitellogenic stage marked the commencement and end of vitellogenesis II, respectively. Yolk granules at various stages of development were seen in the ooplasm from late vitellogenic stage onwards. The spent ovary had an area with resorbing oocytes and empty follicle cells denoting the end of one reproductive cycle and another area with oogonial cells and previtellogenic oocytes indicating the beginning of the next. [ABSTRACT FROM AUTHOR]

Background: The conserved cytokine-induced apoptosis inhibitor-1 ( CIAPIN1) gene has been implicated in several processes, such as apoptosis, cell division, angiogenesis and Fe/S protein biogenesis. In this study, we identified the Drosophila CIAPIN1 homologue ( D-CIAPIN1) and studied its role in ovarian development. Results: We found that D-CIAPIN1 is conserved as it can complement the nonviability of the yeast CIAPIN1-deletion strain. Several D-CIAPIN1 alleles were identified, including one allele in which that codon encoding the highly conserved twin cysteine CX2C motif is mutated, demonstrating for the first time the importance of this motif to protein function. We demonstrated D-CIAPIN1 is an essential gene required for ovarian development. We found that D-CIAPIN1 female mutants are sterile, containing rudimentary ovaries. We noted a decrease in follicle cell numbers in D-CIAPIN1 mutant egg chambers. We further demonstrated that the decrease in follicle cell numbers in D-CIAPIN1 mutants is due to a reduced mitotic index and enhanced cell death. Conclusions: Our study reveals that D-CIAPIN1 is essential for egg chamber development and is required for follicle cell proliferation and survival. Developmental Dynamics 242:731-737, 2013. © 2013 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Nuclei wriggle in the cells of the follicle epithelium of the Drosophila pre-vitellogenic egg primordia. Although similar phenomena have been reported for a number of cultured cell types and some neurons in the zebrafish embryo, the mechanism and importance of the process have remained unexplained. Wriggling involves successive sudden and random minor turns of the nuclei, approximately three twists per minute with roughly 12° per twist, one of which lasts typically for 14 seconds. Wriggling is generated by the growing microtubules seeded throughout the cell cortex, which, while poking the nuclei, buckle and exert 5-40 piconewtons over, 16 seconds. While wriggling, the nuclei drift, 5 μm in a day in the immensely growing follicle cells along the apical-basal axis from the apical to the basal cell region. A .2-fold excess of the microtubules nucleated in the apical cell region, as compared with those seeded in the basal cell cortex, makes the nuclei drift along the apical-basal axis. Nuclear wriggling and positioning appear to be tightly related processes: they cease simultaneously when the nuclei become anchored by the actin cytoskeleton; moreover, colchicine or taxol treatment eliminates both nuclear wriggling and positioning. We propose that the wriggling nuclei reveal a thus far undescribed nuclear positioning mechanism. [ABSTRACT FROM AUTHOR]

Abstract: Estrogen induces oocytes development and vitellogenesis in crustacean by interacting with estrogen receptor (ER) subtypes. In the present study, we detect for the first time the ERα in oocytes and follicle cells and hepatopancreas cells of mysis by immunohistochemistry using a specific ERα antibody. ERα was mainly localized in the nuclei of oocytes and follicle cells, while mainly detected in nuclei of oogonia (OG), previtellogenic oocyte (PR) and endogenous vitellogenic oocyte (EN) at previtellogenic and early vitellogenic stage (I–early III). Follicle cells in all stages of ovary (all vitellogenic stages) showed strong ERα positive reaction, and they were able to gradually move to oocytes during the development of oocytes. In addition, ERα was also localized in the nuclei and cytoplasm of four hepatopancreas cells (including E-, R-, F- and B-cell) in all ovary stages. These findings suggest, for the first time to our knowledge, that there could be a close link between oogenesis, follicle cells, hepatopancreas cells and endocrine regulation, and estrogens might be involved in the regulation of oocytes at early ovarian stage in mysis. [Copyright &y& Elsevier]

Background: The proper balance of autophagy, a lysosome-mediated degradation process, is indispensable for oogenesis in Drosophila. We recently demonstrated that egg development depends on autophagy in the somatic follicle cells (FC), but not in the germline cells (GCs). However, the lack of autophagy only affects oogenesis when FCs are autophagy-deficient but GCs are wild type, indicating that a dysfunctional signaling between soma and germline may be responsible for the oogenesis defects. Thus, autophagy could play an essential role in modulating signal transduction pathways during egg development. Results: Here, we provide further evidence for the necessity of autophagy during oogenesis and demonstrate that autophagy is especially required in subsets of FCs. Generation of autophagy-deficient FCs leads to a wide range of phenotypes that are similar to mutants with defects in the classical cell-cell signaling pathways in the ovary. Interestingly, we observe that loss of autophagy leads to a precocious activation of the Notch pathway in the FCs as monitored by the expression of Cut and Hindsight, two downstream effectors of Notch signaling. Conclusion: Our findings point to an unexpected function for autophagy in the modulation of the Notch signaling pathway during Drosophila oogenesis and suggest a function for autophagy in proper receptor activation. Egg development is affected by an imbalance of autophagy between signal sending (germline) and signal receiving cell (FC), thus the lack of autophagy in the germline is likely to decrease the amount of active ligand and accordingly compensates for increased signaling in autophagy-defective follicle cells. [ABSTRACT FROM AUTHOR]

Introduction: Establishment of distinct follicle cell fates at the early stages of Drosophila oogenesis is crucial for achieving proper morphology of individual egg chambers. In Drosophila oogenesis, Notch-signaling controls proliferation and differentiation of follicular cells, which eventually results in the polarization of the anterior-posterior axis of the oocyte. Here we analyzed the functions of Tribolium Notch-signaling factors during telotrophic oogenesis, which differs fundamentally from the polytrophic ovary of Drosophila. Results: We found Notch-signaling to be required for maintaining the mitotic cycle of somatic follicle cells. Upon Delta RNAi, follicle cells enter endocycle prematurely, which affects egg-chamber formation and patterning. Interestingly, our results indicate that Delta RNAi phenotypes are not solely due to the premature termination of cell proliferation. Therefore, we monitored the terminal/stalk cell precursor lineage by molecular markers. We observed that upon Delta RNAi terminal and stalk cell populations were absent, suggesting that Notch-signaling is also required for the specification of follicle cell populations, including terminal and stalk precursor cells. Conclusions: We demonstrate that with respect to mitotic cycle/endocycle switch Notch-signaling in Tribolium and Drosophila has opposing effects. While in Drosophila a Delta-signal brings about the follicle cells to leave mitosis, Notch-signaling in Triboliumis necessary to retain telotrophic egg-chambers in an "immature" state. In most instances, Notch-signaling is involved in maintaining undifferentiated (or preventing specialized) cell fates. Hence, the role of Notch in Tribolium may reflect the ancestral function of Notch-signaling in insect oogenesis. The functions of Notch-signaling in patterning the follicle cell epithelium suggest that Tribolium oogenesis may - analogous to Drosophila - involve the stepwise determination of different follicle cell populations. Moreover, our results imply that Notch-signaling may contribute at least to some aspects of oocyte polarization and AP axis also in telotrophic oogenesis. [ABSTRACT FROM AUTHOR]

Ovarian follicle cells are the site of yolk protein synthesis in the Ezo abalone Haliotis discus hannai. In this study, histological observations of ovarian follicle cells were conducted in H. discus hannai by immunocytochemical and in situ hybridization methods focusing on their function of yolk protein synthesis. An antibody raised against purified yolk protein (vitellin, Vn) recognized yolk granules in oocytes, and ovarian follicle cells adjacent to the oocytes under yolk accumulation were stained positively with both anti-Vn antibody and an antisense probe for vitellogenin (the precursor for vitellin, Vtg) mRNA. These results indicate that the abalone Vtg gene is transcribed and translated in the ovarian follicle cells. In oocytes in the early phase of yolk accumulation, positive reactions with the antibody appeared first in the stalk part, and the follicle cells adjacent to the stalk were also stained positively. These observations imply local transport of yolk protein from the follicle cells to the oocyte through an extracellular space around the oocyte stalk. Ovarian follicle cells changed their morphological characteristics and reactivity to the antibody in close relation to the stages of the adjacent oocyte, which suggests the presence of functional interactions between follicle cells and oocytes in the course of yolk protein synthesis and accumulation. [ABSTRACT FROM AUTHOR]

Ultrastructural studies of vitellogenesis in oocytes and follicle cells during oogenesis in female Protothaca (Notochione) jedoensis were investigated by histological and transmission electron microscope observations. In early vitellogenic oocytes, combined activities of the Golgi complex, mitochondria and rough endoplasmic reticulum in the cytoplasm are associated with autosynthetic vitellogenesis. Furthermore, at this time, many coated vesicles at the basal region of the oolemma of the oocyte lead to the formation of vesicles through endocytosis in the cytoplasm. Through the formation of the coated pits on oolemma during vitellogenesis, the uptake of extrafollicular precursors (nutritive materials) occurs in coated vesicles by endocytosis. Therefore, it is assumed that these exogenous materials are involved in heterosynthetic vitellogenesis. During late oogenesis, exogenous yolk precursors (yolk granules), lipid droplets and proteinaceous yolk granules are present in the cytoplasm of late vitellogenic oocytes. In mature oocytes, small yolk granules appear intermingled and form large mature yolk granules. Thus, two processes of vitellogenesis occur in oocytes by way of endogenous autosynthesis and exogenous heterosynthesis. The follicle cells attached to the oocytes appear to play an integral role in vitellogenesis in this study. [ABSTRACT FROM AUTHOR]

Abstract: In fly ovaries, the follicular epithelium surrounding germline cells diversifies into several morphologically distinct cell subpopulations. This complex process is crucial for the formation of a regionally complex eggshell and establishment of polarity of the future embryo. Morphogenetic changes accompanying patterning of the follicular epithelium have been best characterized in the model fly, Drosophila melanogaster. Here, we analyze follicular epithelium diversification in the ovaries of Tachypeza nubila, a brachyceran fly closely related to the group Cyclorrhapha, which also includes Drosophila. We provide morphological evidence that in Tachypeza, the diversification process differs from that described in the Drosophila model system in several important respects: (i) follicle cells differentiate into five subpopulations (versus eight in Drosophila); (ii) only one of these subpopulations (i.e. border cells) is migratory (versus four in Drosophila); (iii) the main body follicle cells form a uniform epithelium with no distinct border between follicle cells covering the nurse cell compartment and the oocyte; (iv) chorionic material is deposited not only on the surface of the oocyte but also on the nurse cells; (v) there is no centripetal migration of the follicle cells; (vi) the resulting eggshell is morphologically simple with no regional specializations except for the micropylar apparatus at the anterior pole of the oocyte. Our findings provide novel insights into the evolution of the follicle cell patterning and functioning in dipterans. A critical analysis of these processes in different dipteran groups strongly indicates that in Tachypeza, follicular epithelium diversification follows a distinct pattern, novel for higher dipterans. [Copyright &y& Elsevier]

An essay is presented on the egg formation of lepidopterans. It states that egg formation starts with the early growth and differentiation of female germ cells and then undergoes processes such as morphogenesis, physiology, and biosynthesis. It mentions other developments related on egg formation of lepidopterans including prefollicular development, follicle formation, and follicle dormancy. It adds that lepidopteran ovary had viewed to contribute in the studies of egg formation in all insects.

Senile graying of human hair has been the subject of intense research since ancient times. Reactive oxygen species have been implicated in hair follicle melanocyte apoptosis and DNA damage. Here we show for the first time by FT-Raman spectroscopy vivo that human gray/white scalp hair shafts accumulate hydrogen peroxide (H2O2) in millimolar concentrations. Moreover, we demonstrate almost absent catalase and methionine sulfoxide reductase A and protein expression via immunofluorescence and Western blot in association with a functional loss of methionine sulfoxide (Met-S=O) repair in the entire gray hair follicle. Accordingly, Met-S=O formation of Met residues, including Met 374 in the active site of tyrosinase, the key enzyme in melanogenesis, limits enzyme functionality, as evidenced by FT-Raman spectroscopy, computer simulation, and enzyme kinetics, which leads to gradual loss of hair color. Notably, under in vitro conditions, Met oxidation can be prevented by L-methionine. In summary, our data feed the long-voiced, but insufficiently proven, concept of H2O2-induced oxidative damage in the entire human hair follicle, inclusive of the hair shaft, as a key element in senile hair graying, which does not exclusively affect follicle melanocytes. This new insight could open new strategies for intervention and reversal of the hair graying process. [ABSTRACT FROM AUTHOR]

Abstract: The Balbiani body is an evolutionarily conserved asymmetric aggregate of organelles that is present in early oocytes of all animals examined, including humans. Although first identified more than 150 years ago, genes acting in the assembly of the Balbiani body have not been identified in a vertebrate. Here we show that the bucky ball gene in the zebrafish is required to assemble this universal aggregate of organelles. In the absence of bucky ball the Balbiani body fails to form, and vegetal mRNAs are not localized in oocytes. In contrast, animal pole localized oocyte markers are expanded into vegetal regions in bucky ball mutants, but patterning within the expanded animal pole remains intact. Interestingly, in bucky ball mutants an excessive number of cells within the somatic follicle cell layer surrounding the oocyte develop as micropylar cells, an animal pole specific cell fate. The single micropyle permits sperm to fertilize the egg in zebrafish. In bucky ball mutants, excess micropyles cause polyspermy. Thus bucky ball provides the first genetic access to Balbiani body formation in a vertebrate. We demonstrate that bucky ball functions during early oogenesis to regulate polarity of the oocyte, future egg and embryo. Finally, the expansion of animal identity in oocytes and somatic follicle cells suggests that somatic cell fate and oocyte polarity are interdependent. [Copyright &y& Elsevier]

Abstract: The oocytes of many fish species accumulate high amounts of neutral lipids as a caloric reserve for embryonic and larval development. We propose that lipoprotein lipase (LPL, EC 3.1.1.34) plays an important role in supplying the oocytes with fatty acids and we have cloned its cDNA from the ovary of sea bass, and determined the patterns of LPL activity and LPL mRNA expression in the ovary. The cDNA obtained was 3051 bp long with an open reading frame encoding 518 amino acids. The amino acid sequence has a high similarity and shows similar structural features to LPL of other species. Northern blot analysis revealed LPL expression in adipose tissue and gonads only. LPL activity and LPL mRNA expression in the ovary was very high in fish with a gonadosomatic index (GSI) above 5, coinciding with the appearance of a high number of lipid droplets in the ooplasm. The LPL mRNA expression was localised to the follicle cells surrounding the oocyte. Our results suggest that LPL is likely to play an important role in the incorporation of neutral lipids into the oocytes, and that follicle cells, in addition to participating in steroidogenesis, also may be important in building up oocyte lipid reserves. [Copyright &y& Elsevier]

TSC22D1, which encodes transforming growth factor β-stimulated clone 22 (TSC-22), is thought to be a tumor suppressor because its expression is lost in many glioblastoma, salivary gland, and prostate cancers. TSC-22 is the founding member of the TSC-22/DIP/Bun family of leucine zipper transcription factors; its functions have not been investigated in a multicellular environment. Genetic studies in the model organism Drosophila melanogaster often provide fundamental insights into mechanisms disrupted in carcinogenesis, because of the strong evolutionary conservation of molecular mechanisms between flies and humans. Whereas humans and mice have four TSC-22 domain genes with numerous isoforms, Drosophila has only one TSC-22 domain gene, bunched (bun), which encodes both large and small protein isoforms. Surprisingly, Drosophila Bun proteins promote cellular growth and proliferation in ovarian follicle cells. Loss of both large isoforms has the strongest phenotypes, including increased apoptosis. Cultured 52 cells depleted for large Bun isoforms show increased apoptosis and less frequent cell division, with decreased cell size. Altogether, these data indicate that Drosophila TSC-22/DIP/Bun proteins are necessary for cellular growth, proliferation, and survival both in culture and in an epithelial context. Previous work demonstrated that bun prevents recruitment of epithelial cells to a migratory fate and, thus, maintains epithelial organization. We speculate that reduced TSC22D1 expression generally reduces cellular fitness and only contributes to carcinogenesis in specific tissue environments. [ABSTRACT FROM AUTHOR]

Abstract: Only a few biochemical and molecular studies on yolk proteins (vitellins) have been carried out in mollusks, mainly in bivalves, while information on prosobranch vitellogenesis is still limited. In this study, we cloned a full-length cDNA encoding vitellogenin (Vg) in the Pacific abalone Haliotis discus hannai. The complete Vg cDNA consists of 7753 nucleotides with a long open reading frame encoding 2391 amino acid residues. The deduced primary structure contains the N-terminal amino acid sequences of the 95 kDa and 150 kDa subunits of vitellin of the abalone and shows similarities to Vgs of other mollusk, fish, nematode and coral species. In common with bivalve Vgs, the abalone Vg gene was expressed only in the ovary. In situ hybridization analysis further localized Vg mRNA to the follicle cells in the ovary. We conclude that the follicle cells are the site of Vg synthesis in H. discus hannai. [Copyright &y& Elsevier]

Abstract: Two proteins from the eggshell of Rhodnius prolixus were isolated, characterized and named Rp30 and Rp45 according to their molecular masses. Purified proteins were used to obtain specific antiserum which was later used for immunolocalization. The antiserum against Rp30 and Rp45 detected their presence inside the follicle cells, their secretion and their association with oocyte microvilli. Both proteins are expressed during the final stage of vitellogenesis, preserved during embryogenesis and discarded together with the eggshell. The amino terminals were sequenced and both proteins were further cloned using degenerated primers. The amino acid sequences appear to have a tripartite arrangement with a highly conserved central domain which presents a repetitive motif of valine–proline–valine (VPV) at intervals of 15 amino acid residues. Their amino acid sequence showed no similarity to any known eggshell protein. The expression of these proteins was also investigated; the results demonstrated that this occurred strictly in choriogenic follicles. Antifungal activity against Aspergillus niger was found to be associated with Rp45 but not with Rp30. A. niger exposed to Rp45 protein induced growth inhibition and several morphological changes such as large vacuoles, swollen mitochondria, multi-lamellar structures and a disorganized cell wall as demonstrated by electron microscopy analysis. [Copyright &y& Elsevier]

Abstract: STAM (signal-transducing adaptor molecule) is a protein highly conserved from yeast to mammals. In Drosophila melanogaster the basic molecular architecture of the protein is comprised of a N-terminal VHS domain, an ubiquitin-interacting motif and a central Src homology-3 domain. In this paper we examine the expression pattern of the stam gene and the localisation of the STAM protein during D. melanogaster oogenesis. Its transcript is present throughout egg chamber development in all germ-line cells, including the oocyte. dSTAM is firstly detected in germarial region 2, where the protein is present in the newly formed germ-line cysts and is mainly accumulated into the oocyte. As oogenesis proceeds, dSTAM is enriched in the perinuclear region of the nurse cells and is also found in the somatic polar follicular cells. In the oocyte, the protein is more abundant posteriorly and becomes restricted to the posterior pole just before disappearing at stage 10b. We show that dSTAM localisation is unaffected in the oocyte of grk mutant egg chambers, indicating that it is not dependent on the polarity of the microtubule network. In contrast, dSTAM distribution is remarkably altered in cup mutant oocytes where the protein accumulates in a round central spot and never reaches the posterior pole. [Copyright &y& Elsevier]

Abstract: A cDNA encoding the Pacific oyster, Crassostrea gigas, estrogen receptor (cgER) was cloned using degenerate PCR primers. The open reading frame predicted 485 amino acid residues. Comparisons of the amino acid sequence of cgER with other mollusk ERs show high similarities of the C domain (95–97%), and the E domain (56–66%). The amino acid sequence of the C domain of cgER shows 86 and 89% identity with the respective sequences of human ER-α and ER-β. The amino acid sequence of the E domain of cgER shows 45% identity with those of human ER-α and ER-β. The phylogenetic analysis indicated that the cgER is an ortholog of the other mollusk ERs. In the C domain, the positions of cysteine residues and other residues around them that constitute the two zinc finger motifs and the P-box are conserved. The cgER mRNA was expressed in various tissues including the ovary. Reporter gene assay revealed that cgER is unresponsive to estrogen. This result is similar to those of other mollusk ERs. ER immunoreactivity was localized mainly in the nuclei of follicle cells, the site of vitellogenin synthesis, and in oocytes. This result suggests that cgER could work as a nuclear receptor. [Copyright &y& Elsevier]

Abstract: Heptachlor, a chlorinated hydrocarbon pesticide, suppresses the production of progesterone and estradiol in the female rat in vivo or in isolated ovaries in vitro. In this study the effect of heptachlor on steroid hormone production by isolated rat luteal and follicular cells, in the presence of two precursor hormones was investigated. Ovaries were isolated from anesthetized mature normocyclic virgin rats (3 to 4 months old), under sterile conditions. Corpora lutea and follicles were microscopically dissected out and separately enzymatically dispersed with collagenase at 37 °C. Viable cells collected after centrifugation were used at a concentration of approximately 2.5×105 cells/10 mL. Both luteal and follicular cell preparations were separately incubated overnight (15 h) at 37 °C in the presence of pregnenolone (P5) and androstenedione (A4) at a concentration of 6.0 nmol/L each, and heptachlor at either 0.12 μg/mL (low dose) or 1.20 μg/mL (high dose) (test cells) or in the absence of heptachlor (control cells). At the end of the incubations, progesterone and estradiol 17β levels were analyzed in the incubation media. The results indicate that heptachlor significantly suppressed the production of both progesterone and estradiol in both cell types in a dose related manner even in the presence of A4 and P5 as precursor hormones (P <0.05). [Copyright &y& Elsevier]