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The leading cause of infertility in men and women is quantitative and qualitative defects in human germ-cell (oocyte and sperm) development. Yet, it has not been possible to examine the [...]

The Retinoblastoma (Rb) protein is a conserved repressor of cell proliferation. In animals and plants, deregulation of Rb protein causes hyperproliferation and perturbs cell differentiation to various degrees. However, the primary developmental impact of the loss of Rb protein has remained unclear. In this study we investigated the direct consequences of Rb protein knockout in the Arabidopsis male germline using cytological and molecular markers. The Arabidopsis germ line derives from the unequal division of the microspore, producing a small germ cell and a large terminally differentiated vegetative cell. A single division of the germ cell produces the 2 sperm cells. We observed that the loss of Rb protein does not have a major impact on microspore division but causes limited hyperproliferation of the vegetative cell and, to a lesser degree, of the sperm cells. In addition, cell fate is perturbed in a fraction of Rb-defective vegetative cells. These defects are rescued by preventing cell proliferation arising from down-regulation of cyclin-dependent kinase A1. Our results indicate that hyperproliferation caused by the loss of Rb protein prevents or delays cell determination during plant male gametogenesis, providing further evidence for a direct link between fate determination and cell proliferation. male germline | pollen | cell cycle

Author(s): H. Diego Folco [1]; Venkata R. Chalamcharla [1]; Tomoyasu Sugiyama [1]; Gobi Thillainadesan [1]; Martin Zofall [1]; Vanivilasini Balachandran [1]; Jothy Dhakshnamoorthy [1]; Takeshi Mizuguchi [1]; Shiv I. S. [...]

We applied independent species concepts to clarify the phylogeographic structure of the ascidian Ciona intestinalis, a powerful model system in chordate biology and for comparative genomic studies. Intensive research with this marine invertebrate is based on the assumption that natural populations globally belong to a single species. Therefore, understanding the true taxonomic classification may have implications for experimental design and data management. Phylogenies inferred from mitochondrial and nuclear DNA markers accredit the existence of two cryptic species: C. intestinalis sp. A, genetically homogeneous, distributed in the Mediterranean, northeast Atlantic, and Pacific, and C. intestinalis sp. B, geographically structured and encountered in the North Atlantic. Species-level divergence is further entailed by cross-breeding estimates. C. intestinalis A and B from allopatric populations cross-fertilize, but hybrids remain infertile because of defective gametogenesis. Although anatomy illustrates an overall interspecific similarity lacking in diagnostic features, we provide consistent tools for in-field and in-laboratory species discrimination. Finding of two cryptic taxa in C. intestinalis raises interest in a new tunicate genome as a gateway to studies in speciation and ecological adaptation of chordates. anatomy | Ciona intestinalis | cryptic species I gamete compatibility | phylogenetics

We report a cross-species expression profiling analysis of the human, mouse, and rat male meiotic transcriptional program, using enriched germ cell populations, whole gonads, and high-density oligonucleotide microarrays (GeneChips). Among 35% of the protein-coding genes present in rodent and human genomes that were found to be differentially expressed between germ cells and somatic controls, a key group of 357 conserved core loci was identified that displays highly similar meiotic and postmeiotic patterns of transcriptional induction across all three species. Genes known to be important for sexual reproduction are significantly enriched among differentially expressed core loci and a smaller group of conserved genes not detected in 17 nontesticular somatic tissues, correlating transcriptional activation and essential function in the male germ line. Some genes implicated in the etiology of cancer are found to be strongly transcribed in testis, suggesting that these genes may play unexpected roles in sexual reproduction. Expression profiling data further identified numerous conserved genes of biological and clinical interest previously unassociated with the mammalian male germ line. bioinformatics | spermatogenesis | transcriptome

In Anopheles gambiae, the vector for the most deadly malaria parasite Plasmodium falciparum, xanthurenic acid (XA) plays a key role in parasite gametogenesis and fertility. In mosquitoes, XA is produced by transamination of 3-hydroxykynurenine (3-HK), a reaction that represents the main route to prevent the accumulation of the potentially toxic 3-HK excess. Interfering with XA metabolism in A. gambiae therefore appears an attractive avenue for the development of malaria transmission-blocking drugs and insecticides. We have determined the crystal structure of A. gambiae 3-HK transaminase in its pyridoxal 5'-phosphate form and in complex with a newly synthesized competitive enzyme inhibitor. Structural inspection of the enzyme active site reveals the key molecular determinants for ligand recognition and catalysis. Major contributions toward inhibitor binding are provided by a salt bridge between the inhibitor carboxylate and Arg-356 and by a remarkable hydrogen bond network involving the anthranilic moiety of the inhibitor and backbone atoms of residues Gly-25 and Asn-44. This study may be useful for the structurebased design of specific enzyme inhibitors of potential interest as antimalarial agents. kynurenine pathway | xanthurenic acid

Chromatin-remodeling factors regulate the establishment of transcriptional programs during plant development. Although 42 genes encoding members of the SWI2/SNF2 family have been identified in Arabidopsis thaliana, imitation of switch proteins | megagametophyte | RNA interference | seed development | functional megaspore

Byline: Hilal Ilarslan (1), Harry T. Horner (2), Reid G. Palmer (3) Keywords: Confocal laser scanning microscopy Glycine max Megagametophyte Ovule abortion Partial sterility Abstract: Megagametogenesis of soybean, Glycine max (L.) Merr., cultivars Clark and Clark k2, and F1 hybrid of Clark (female parent) crossed with Clark k2 (male parent) were studied using stereo light microscopy and confocal scanning laser microscopy. Reproductive development in Clark and Clark k2 plants was compared to F1 hybrid plants. In mature pods, 6.4% of the ovules of Clark, 8.1% of the ovules of Clark k2, and 41.4% of the ovules of F1 hybrid plants were aborted. This female partial sterility was due to incomplete megagametophyte development: undeveloped polar nuclei--or developed but not in a position for fertilization increased megagametophyte wall thickness abnormal shape and/or premature degeneration of synergids and intact synergids throughout the life of the ovule egg cell not well-developed or absent and megagametophyte remaining uninucleate. Each of these abnormalities contributed to either lack of double fertilization or early megagametophyte abortion. Author Affiliation: (1) Department of Biology, Ankara University, 06100, Tandogan, Ankara, Turkey (2) Department of Botany and Bessey Microscopy Facility, Iowa State University, IA 50011-1020, Ames, USA (3) USDA ARS-CICGR and Departments of Agronomy and of Zoology/Genetics, G301 Agronomy, Iowa State University, IA 50011-1010, Ames, USA Article History: Received Date: 29/07/2002 Accepted Date: 20/12/2002 Article note: Electronic Publication

Although fertilization has been studied for more than a century, the cell surface proteins mediating the process are only now becoming known. Gamete interaction in animals appears to be molecularly complex. Although it is difficult to generalize at present, diversity of structure may be a recurring theme in the evolution of fertilization proteins. Examples of rapid evolution of fertilization proteins by positive selection are known, and concerted evolution can influence the differentiation of gamete recognition proteins between closely related species., Sexual reproduction, defined here as the fusion of two haploid cells during fertilization to form a diploid zygote, occurs in almost all eukaryotes. Fertilization is the bridge between generations. Although [...]

The role of A-myb in spermatogenesis and mammary gland development was established by utilizing genomic fragments cloned from a-myb homozygous mice. Results revealed that a-myb gene spliced mice develop to full term but exhibited defects in growth and male infertility due to arrested spermeiogenesis at the pachytene phase. Female adult a-myb gene spliced mice show underdeveloped lactating breast tissues.

In addition to the five 70-kDa heat shock proteins (HSP70) common to germ cells and somatic tissues of mammals, spermatogenic cells synthesize HSP70-2 during meiosis. To determine if this unique stress protein has a critical role in meiosis, we used gene-targeting techniques to disrupt Hsp70-2 in mice. Male mice homozygous for the mutant allele (Hsp70-[2.sup.- / -]) did not synthesize HSP70-2, lacked postmeiotic spermatids and mature sperm, and were infertile. However, neither meiosis nor fertility was affected in female Hsp70-[2.sup.- / -] mice. We previously found that HSP70-2 is associated with synaptonemai complexes in the nucleus of meiotic spermatocytes from mice and hamsters. While synaptonemal complexes assembled in Hsp70-[2.sup.- / -] spermatocytes, structural abnormalities became apparent in these cells by late prophase, and development rarely progressed to the meiotic divisions. Furthermore, analysis of nuclei and genomic DNA indicated that the failure of meiosis in Hsp70-[2.sup.- / -] mice was coincident with a dramatic increase in spermatocyte apo-ptosis. These results suggest that HSP70-2 participates in synaptonemal complex function during meiosis in male germ cells and is linked to mechanisms that inhibit apoptosis.

The cDNA coding for the Chlamydomonas reinhardtii gamete lytic enzyme (GLE) was isolated and characterized. Sequence analyses showed that GLE is synthesized in a preproenzyme form. Sequence comparisons further localized the putative autocatalytic activation, zinc-binding and calcium-binding sites. Northern blot analyses showed that GLE mRNA expression is increased during the vegetative cell cycle and gametic differentiation. The results suggest that vegetative cells synthesize and store proGLE which is released upon sonification. During gametogenesis, this is shifted to a locale that allows release by homogenization, affecting a conversion to GLE.

An analysis of the basic properties of Linkage Disequilibrium (LD), variation in LD among species, and the application of LD in the dissection of quantitative traits is given. The patterns of LD are influenced by the mating system of the species, and phenomena such as population structure and recombination hot spots.

In analyzing the transcriptional networks that regulate development, one ideally would like to determine whether a particular transcription factor binds directly to a candidate target promoter inside the living embryo. Properties of the Caenorhabditis elegans elt-2 gene, which encodes a gut-specific GATA factor, have allowed us to develop such a method. We previously have shown, by means of ectopic expression studies, that elt-2 regulates its own promoter. To test whether this autoregulation is direct, we fused green fluorescent protein (GFP) close to the C terminus of elt-2 in a construct that contains the full elt-2 promoter and the full elt-2 zinc finger DNA binding domain; the construct is expressed correctly (i.e., only in the gut lineage) and is able to rescue the lethality of an elt-2 null mutant. Multicopy transgenic arrays of this rescuing elt-2::GFP construct were integrated into the genome and transgenic embryos were examined when the developing gut has 4-8 cells; the majority of these embryonic gut nuclei show two discrete intense foci of fluorescence. We interpret these fluorescent foci as the result of ELT-2::GFP binding directly to its own promoter within nuclei of the developing gut lineage. Numerous control experiments, both genetic and biochemical, all support this conclusion and support the specificity of the binding. The approach should be applicable to studying other transcription factors binding target promoters, all within the living C. elegans embryo.

Mammals inherit two complete sets of chromosomes from their parents and thus two copies of every autosomal gene. Normally both copies are expressed, but, in a minority of cases, a [...]