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Author(s): M. Bettencourt-Dias (corresponding author) [1]; R. Giet [1, 2]; R. Sinka [1]; A. Mazumdar [1]; W. G. Lock [1]; F. Balloux [1]; P. J. Zafiropoulos [1]; S. Yamaguchi [3]; [...]

Adult newt cardiomyocytes, in contrast to their mammalian counterparts, can proliferate after injury and contribute to the functional regeneration of the heart. In order to understand the mechanisms underlying this plasticity we performed longitudinal studies on single cardiomyocytes in culture. We find that the majority of cardiomyocytes can enter S phase, a process that occurs in response to serum-activated pathways and is dependent on the phosphorylation of the retinoblastoma protein. However, more than half of these cells stably arrest at either entry to mitosis or during cytokinesis, thus resembling the behaviour observed in mammalian cardiomyocytes. Approximately a third of the cells progress through mitosis and may enter successive cell divisions. When cardiomyocytes divided more than once, the proliferative behaviour of sister cells was significantly correlated, in terms of whether they underwent a subsequent cell cycle, and if so, the duration of that cycle. These observations suggest a mechanism whereby newt heart regeneration depends on the retention of proliferative potential in a subset of cardiomyocytes. The regulation of the remaining newt cardiomyocytes is similar to that described for their mammalian counterparts, as they arrest during mitosis or cytokinesis. Understanding the nature of this block and why it arises in some but not other newt cardiomyocytes may lead to an augmentation of the regenerative potential in the mammalian heart. [ABSTRACT FROM AUTHOR]

Ciliary defects cause several ciliopathies, some of which have late onset, suggesting cilia are actively maintained. Still, we have a poor understanding of the mechanisms underlying their maintenance. Here, we show Drosophila melanogaste r IFT88 ( Dm IFT88/nompB) continues to move along fully formed sensory cilia. We further identify Inactive, a TRPV channel subunit involved in Drosophila hearing and negative-gravitaxis behaviour, and a yet uncharacterised Drosophila Guanylyl Cyclase 2d ( Dm Gucy2d/CG34357) as Dm IFT88 cargoes. We also show Dm IFT88 binding to the cyclase´s intracellular part, which is evolutionarily conserved and mutated in several degenerative retinal diseases, is important for the ciliary localisation of Dm Gucy2d. Finally, acute knockdown of both Dm IFT88 and Dm Gucy2d in ciliated neurons of adult flies caused defects in the maintenance of cilium function, impairing hearing and negative-gravitaxis behaviour, but did not significantly affect ciliary ultrastructure. We conclude that the sensory ciliary function underlying hearing in the adult fly requires an active maintenance program which involves Dm IFT88 and at least two of its signalling transmembrane cargoes, Dm Gucy2d and Inactive., (© 2024 Werner et al.)

Centrioles are part of centrosomes and cilia, which are microtubule organising centres (MTOC) with diverse functions. Despite their stability, centrioles can disappear during differentiation, such as in oocytes, but little is known about the regulation of their structural integrity. Our previous research revealed that the pericentriolar material (PCM) that surrounds centrioles and its recruiter, Polo kinase, are downregulated in oogenesis and sufficient for maintaining both centrosome structural integrity and MTOC activity. We now show that the expression of specific components of the centriole cartwheel and wall, including ANA1/CEP295, is essential for maintaining centrosome integrity. We find that Polo kinase requires ANA1 to promote centriole stability in cultured cells and eggs. In addition, ANA1 expression prevents the loss of centrioles observed upon PCM-downregulation. However, the centrioles maintained by overexpressing and tethering ANA1 are inactive, unlike the MTOCs observed upon tethering Polo kinase. These findings demonstrate that several centriole components are needed to maintain centrosome structure. Our study also highlights that centrioles are more dynamic than previously believed, with their structural stability relying on the continuous expression of multiple components., (© 2024. The Author(s).)

Polo-like kinase 4 (Plk4), the major regulator of centriole biogenesis, has emerged as a putative therapeutic target in cancer due to its abnormal expression in human carcinomas, leading to centrosome number deregulation, mitotic defects and chromosomal instability. Moreover, Plk4 deregulation promotes tumor growth and metastasis in mouse models and is significantly associated with poor patient prognosis. Here, we further investigate the role of Plk4 in carcinogenesis and show that its overexpression significantly potentiates resistance to cell death by anoikis of nontumorigenic p53 knock-out (p53KO) mammary epithelial cells. Importantly, this effect is independent of Plk4's role in centrosome biogenesis, suggesting that this kinase has additional cellular functions. Interestingly, the Plk4-induced anoikis resistance is associated with the induction of a stable hybrid epithelial-mesenchymal phenotype and is partially dependent on P-cadherin upregulation. Furthermore, we found that the conditioned media of Plk4-induced p53KO mammary epithelial cells also induces anoikis resistance of breast cancer cells in a paracrine way, being also partially dependent on soluble P-cadherin secretion. Our work shows, for the first time, that high expression levels of Plk4 induce anoikis resistance of both mammary epithelial cells with p53KO background, as well as of breast cancer cells exposed to their secretome, which is partially mediated through P-cadherin upregulation. These results reinforce the idea that Plk4, independently of its role in centrosome biogenesis, functions as an oncogene, by impacting the tumor microenvironment to promote malignancy., (© 2023. The Author(s).)

The actin motor protein myosin VI is a multivalent protein with diverse functions. Here, we identified and characterised a myosin VI ubiquitous interactor, the oral-facial-digital syndrome 1 (OFD1) protein, whose mutations cause malformations of the face, oral cavity, digits and polycystic kidney disease. We found that myosin VI regulates the localisation of OFD1 at the centrioles and, as a consequence, the recruitment of the distal appendage protein Cep164. Myosin VI depletion in non-tumoural cell lines causes an aberrant localisation of OFD1 along the centriolar walls, which is due to a reduction in the OFD1 mobile fraction. Finally, loss of myosin VI triggers a severe defect in ciliogenesis that could be, at least partially, ascribed to an impairment in the autophagic removal of OFD1 from satellites. Altogether, our results highlight an unprecedent layer of regulation of OFD1 and a pivotal role of myosin VI in coordinating the formation of the distal appendages and primary cilium with important implications for the genetic disorders known as ciliopathies., (© 2021 The Authors. Published under the terms of the CC BY NC ND 4.0 license.)

Centrioles are structurally conserved organelles, composing both centrosomes and cilia. In animal cycling cells, centrioles often form through a highly characterized process termed canonical duplication. However, a large diversity of eukaryotes assemble centrioles de novo through uncharacterized pathways. This unexplored diversity is key to understanding centriole assembly mechanisms and how they evolved to assist specific cellular functions. Here, we show that, during spermatogenesis of the bryophyte Physcomitrium patens, centrioles are born as a co-axially oriented centriole pair united by a cartwheel. Interestingly, we observe that these centrioles are twisted in opposite orientations. Microtubules emanate from the bicentrioles, which localize to the spindle poles during cell division. After their separation, the two resulting sister centrioles mature asymmetrically, elongating specific microtubule triplets and a naked cartwheel. Subsequently, two motile cilia are assembled that appear to alternate between different motility patterns. We further show that centriolar components SAS6, Bld10, and POC1, which are conserved across eukaryotes, are expressed during spermatogenesis and required for this de novo biogenesis pathway. Our work supports a scenario where centriole biogenesis, while driven by conserved molecular modules, is more diverse than previously thought., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021. Published by Elsevier Inc.)

The centrosome is a main orchestrator of the animal cellular microtubule cytoskeleton. Dissecting its structure and assembly mechanisms has been a goal of cell biologists for over a century. In the last two decades, a good understanding of the molecular constituents of centrosomes has been achieved. Moreover, recent breakthroughs in electron and light microscopy techniques have enabled the inspection of the centrosome and the mapping of its components with unprecedented detail. However, we now need a profound and dynamic understanding of how these constituents interact in space and time. Here, we review the latest findings on the structural and molecular architecture of the centrosome and how its biogenesis is regulated, highlighting how biophysical techniques and principles as well as quantitative modeling are changing our understanding of this enigmatic cellular organelle.

The presence of extra centrioles, termed centrosome amplification, is a hallmark of cancer. The distribution of centriole numbers within a cancer cell population appears to be at an equilibrium maintained by centriole overproduction and selection, reminiscent of mutation-selection balance. It is unknown to date if the interaction between centriole overproduction and selection can quantitatively explain the intra- and inter-population heterogeneity in centriole numbers. Here, we define mutation-selection-like models and employ a model selection approach to infer patterns of centriole overproduction and selection in a diverse panel of human cell lines. Surprisingly, we infer strong and uniform selection against any number of extra centrioles in most cell lines. Finally we assess the accuracy and precision of our inference method and find that it increases non-linearly as a function of the number of sampled cells. We discuss the biological implications of our results and how our methodology can inform future experiments., Competing Interests: The authors have declared that no competing interests exist.

How cells control the numbers of subcellular components is a fundamental question in biology. Given that biosynthetic processes are fundamentally stochastic it is utterly puzzling that some structures display no copy number variation within a cell population. Centriole biogenesis, with each centriole being duplicated once and only once per cell cycle, stands out due to its remarkable fidelity. This is a highly controlled process, which depends on low-abundance rate-limiting factors. How can exactly one centriole copy be produced given the variation in the concentration of these key factors? Hitherto, tentative explanations of this control evoked lateral inhibition- or phase separation-like mechanisms emerging from the dynamics of these rate-limiting factors but how strict centriole number is regulated remains unsolved. Here, a novel solution to centriole copy number control is proposed based on the assembly of a centriolar scaffold, the cartwheel. We assume that cartwheel building blocks accumulate around the mother centriole at supercritical concentrations, sufficient to assemble one or more cartwheels. Our key postulate is that once the first cartwheel is formed it continues to elongate by stacking the intermediate building blocks that would otherwise form supernumerary cartwheels. Using stochastic models and simulations, we show that this mechanism may ensure formation of one and only one cartwheel robustly over a wide range of parameter values. By comparison to alternative models, we conclude that the distinctive signatures of this novel mechanism are an increasing assembly time with cartwheel numbers and the translation of stochasticity in building block concentrations into variation in cartwheel numbers or length., Competing Interests: The authors have declared that no competing interests exist.

Centrioles form centrosomes and cilia. In most proliferating cells, centrioles assemble through canonical duplication, which is spatially, temporally, and numerically regulated by the cell cycle and the presence of mature centrioles. However, in certain cell types, centrioles assemble de novo, yet by poorly understood mechanisms. Herein, we established a controlled system to investigate de novo centriole biogenesis, using Drosophila melanogaster egg explants overexpressing Polo-like kinase 4 (Plk4), a trigger for centriole biogenesis. We show that at a high Plk4 concentration, centrioles form de novo, mature, and duplicate, independently of cell cycle progression and of the presence of other centrioles. Plk4 concentration determines the temporal onset of centriole assembly. Moreover, our results suggest that distinct biochemical kinetics regulate de novo and canonical biogenesis. Finally, we investigated which other factors modulate de novo centriole assembly and found that proteins of the pericentriolar material (PCM), and in particular γ-tubulin, promote biogenesis, likely by locally concentrating critical components., (© 2021 Nabais et al.)

Tuberous sclerosis complex (TSC) is a neurogenetic disorder that leads to elevated mechanistic targeting of rapamycin complex 1 (mTORC1) activity. Cilia can be affected by mTORC1 signaling, and ciliary deficits are associated with neurodevelopmental disorders. Here, we examine whether neuronal cilia are affected in TSC. We show that cortical tubers from TSC patients and mutant mouse brains have fewer cilia. Using high-content image-based assays, we demonstrate that mTORC1 activity inversely correlates with ciliation in TSC1/2-deficient neurons. To investigate the mechanistic relationship between mTORC1 and cilia, we perform a phenotypic screen for mTORC1 inhibitors with TSC1/2-deficient neurons. We identify inhibitors of the heat shock protein 90 (Hsp90) that suppress mTORC1 through regulation of phosphatidylinositol 3-kinase (PI3K)/Akt signaling. Pharmacological inhibition of Hsp90 rescues ciliation through downregulation of Hsp27. Our study uncovers the heat-shock machinery as a druggable signaling node to restore mTORC1 activity and cilia due to loss of TSC1/2, and it provides broadly applicable platforms for studying TSC-related neuronal dysfunction., Competing Interests: Declaration of Interests M.S. reports grant support from Novartis, Roche, Pfizer, Ipsen, LAM Therapeutics, Astellas, Bridgebio, and Quadrant Biosciences unrelated to this project. He has served on Scientific Advisory Boards for Sage, Roche, Aeovian, Celgene, and Takeda. M.S. and A.D.N. have a patent pending on this work. R.J.K. is a current employee and shareholder of Biogen. The other authors declare no competing interests., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Pimento-Marques and Bettencourt-Dias discuss the composition, assembly and function of pericentriolar material - the proteinaceous material that surrounds the centrioles and forms the centrosome, the main microtubule organizing center found in animal cells., (Copyright © 2020 Elsevier Inc. All rights reserved.)

In this Primer, Nabais et al. discuss the evolution of the structure and function of centrioles and basal bodies, describe conserved centriole assembly features and the diversity in centriole architecture across eukaryotes, and highlight important outstanding evolutionary questions concerning centriole assembly., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Centrioles assemble centrosomes and cilia/flagella, which are microtubule-based structures with key roles in cell division, polarity, motility, and signaling. Centriole biogenesis is a tightly regulated process, and deregulation of centriole numbers and structure can have dramatic consequences for cellular function and integrity. However, their small size poses a challenge to study them. Here, we describe protocols that allow the identification and assessment of true centrioles and that provide straightforward strategies to study the role of new candidate proteins in centriole duplication and elongation.

The centrosome is composed of two centrioles surrounded by a microtubule-nucleating pericentriolar material (PCM). Although centrioles are known to regulate PCM assembly, it is less known whether and how the PCM contributes to centriole assembly. Here we investigate the interaction between centriole components and the PCM by taking advantage of fission yeast, which has a centriole-free, PCM-containing centrosome, the SPB. Surprisingly, we observed that several ectopically-expressed animal centriole components such as SAS-6 are recruited to the SPB. We revealed that a conserved PCM component, Pcp1/pericentrin, interacts with and recruits SAS-6. This interaction is conserved and important for centriole assembly, particularly its elongation. We further explored how yeasts kept this interaction even after centriole loss and showed that the conserved calmodulin-binding region of Pcp1/pericentrin is critical for SAS-6 interaction. Our work suggests that the PCM not only recruits and concentrates microtubule-nucleators, but also the centriole assembly machinery, promoting biogenesis close by., Competing Interests: DI, SZ, SJ, PD, JS, ZC, JP, MF, MB No competing interests declared, (© 2019, Ito et al.)

Centrosome amplification (CA) is a common feature of human tumours and a promising target for cancer therapy. However, CA's pan-cancer prevalence, molecular role in tumourigenesis and therapeutic value in the clinical setting are still largely unexplored. Here, we used a transcriptomic signature (CA20) to characterise the landscape of CA-associated gene expression in 9,721 tumours from The Cancer Genome Atlas (TCGA). CA20 is upregulated in cancer and associated with distinct clinical and molecular features of breast cancer, consistently with our experimental CA quantification in patient samples. Moreover, we show that CA20 upregulation is positively associated with genomic instability, alteration of specific chromosomal arms and C>T mutations, and we propose novel molecular players associated with CA in cancer. Finally, high CA20 is associated with poor prognosis and, by integrating drug sensitivity with drug perturbation profiles in cell lines, we identify candidate compounds for selectively targeting cancer cells exhibiting transcriptomic evidence for CA., Competing Interests: The authors have declared that no competing interests exist.

The centrosome is an important microtubule-organising centre (MTOC) in animal cells. It consists of two barrel-shaped structures, the centrioles, surrounded by the pericentriolar material (PCM), which nucleates microtubules. Centrosomes can form close to an existing structure (canonical duplication) or de novo How centrosomes form de novo is not known. The master driver of centrosome biogenesis, PLK4, is critical for the recruitment of several centriole components. Here, we investigate the beginning of centrosome biogenesis, taking advantage of Xenopus egg extracts, where PLK4 can induce de novo MTOC formation ( Eckerdt et al., 2011; Zitouni et al., 2016). Surprisingly, we observe that in vitro , PLK4 can self-assemble into condensates that recruit α- and β-tubulins. In Xenopus extracts, PLK4 assemblies additionally recruit STIL, a substrate of PLK4, and the microtubule nucleator γ-tubulin, forming acentriolar MTOCs de novo The assembly of these robust microtubule asters is independent of dynein, similar to what is found for centrosomes. We suggest a new mechanism of action for PLK4, where it forms a self-organising catalytic scaffold that recruits centriole components, PCM factors and α- and β-tubulins, leading to MTOC formation.This article has an associated First Person interview with the first author of the paper., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2018. Published by The Company of Biologists Ltd.)

Cilia are evolutionarily conserved structures with many sensory and motility-related functions. The ciliary base, composed of the basal body and the transition zone, is critical for cilia assembly and function, but its contribution to cilia diversity remains unknown. Hence, we generated a high-resolution structural and biochemical atlas of the ciliary base of four functionally distinct neuronal and sperm cilia types within an organism, Drosophila melanogaster. We uncovered a common scaffold and diverse structures associated with different localization of 15 evolutionarily conserved components. Furthermore, CEP290 (also known as NPHP6) is involved in the formation of highly diverse transition zone links. In addition, the cartwheel components SAS6 and ANA2 (also known as STIL) have an underappreciated role in basal body elongation, which depends on BLD10 (also known as CEP135). The differential expression of these cartwheel components contributes to diversity in basal body length. Our results offer a plausible explanation to how mutations in conserved ciliary base components lead to tissue-specific diseases.

The centrosome is the major microtubule organizing centre (MTOC) in animal cells. The canonical centrosome is composed of two centrioles surrounded by a pericentriolar matrix (PCM). In contrast, yeasts and amoebozoa have lost centrioles and possess acentriolar centrosomes—called the spindle pole body (SPB) and the nucleus-associated body (NAB), respectively. Despite the difference in their structures, centriolar centrosomes and SPBs not only share components but also common biogenesis regulators. In this review, we focus on the SPB and speculate how its structures evolved from the ancestral centrosome. Phylogenetic distribution of molecular components suggests that yeasts gained specific SPB components upon loss of centrioles but maintained PCM components associated with the structure. It is possible that the PCM structure remained even after centrosome remodelling due to its indispensable function to nucleate microtubules. We propose that the yeast SPB has been formed by a step-wise process; (1) an SPB-like precursor structure appeared on the ancestral centriolar centrosome; (2) it interacted with the PCM and the nuclear envelope; and (3) it replaced the roles of centrioles. Acentriolar centrosomes should continue to be a great model to understand how centrosomes evolved and how centrosome biogenesis is regulated.

Centrosome abnormalities are a typical hallmark of human cancers. However, the origin and dynamics of such abnormalities in human cancer are not known. In this study, we examined centrosomes in Barrett's esophagus tumorigenesis, a well-characterized multistep pathway of progression, from the premalignant condition to the metastatic disease. This human cancer model allows the study of sequential steps of progression within the same patient and has representative cell lines from all stages of disease. Remarkably, centrosome amplification was detected as early as the premalignant condition and was significantly expanded in dysplasia. It was then present throughout malignant transformation both in adenocarcinoma and metastasis. The early expansion of centrosome amplification correlated with and was dependent on loss of function of the tumor suppressor p53 both through loss of wild-type expression and hotspot mutations. Our work shows that centrosome amplification in human tumorigenesis can occur before transformation, being repressed by p53. These findings suggest centrosome amplification in humans can contribute to tumor initiation and progression., (© 2018 Lopes et al.)

Centrosomes are the major microtubule organising centres of animal cells. Deregulation in their number occurs in cancer and was shown to trigger tumorigenesis in mice. However, the incidence, consequence and origins of this abnormality are poorly understood. Here, we screened the NCI-60 panel of human cancer cell lines to systematically analyse centriole number and structure. Our screen shows that centriole amplification is widespread in cancer cell lines and highly prevalent in aggressive breast carcinomas. Moreover, we identify another recurrent feature of cancer cells: centriole size deregulation. Further experiments demonstrate that severe centriole over-elongation can promote amplification through both centriole fragmentation and ectopic procentriole formation. Furthermore, we show that overly long centrioles form over-active centrosomes that nucleate more microtubules, a known cause of invasiveness, and perturb chromosome segregation. Our screen establishes centriole amplification and size deregulation as recurrent features of cancer cells and identifies novel causes and consequences of those abnormalities.

The centriole is a multifunctional structure that organizes centrosomes and cilia and is important for cell signaling, cell cycle progression, polarity, and motility. Defects in centriole number and structure are associated with human diseases including cancer and ciliopathies. Discovery of the centriole dates back to the 19th century. However, recent advances in genetic and biochemical tools, development of high-resolution microscopy, and identification of centriole components have accelerated our understanding of its assembly, function, evolution, and its role in human disease. The centriole is an evolutionarily conserved structure built from highly conserved proteins and is present in all branches of the eukaryotic tree of life. However, centriole number, size, and organization varies among different organisms and even cell types within a single organism, reflecting its cell type-specialized functions. In this review, we provide an overview of our current understanding of centriole biogenesis and how variations around the same theme generate alternatives for centriole formation and function., (© 2018 Loncarek and Bettencourt Dias.)

Centrosomes and cilia are present in organisms from all branches of the eukaryotic tree of life. These structures are composed of microtubules and various other proteins, and are required for a plethora of cell processes such as structuring the cytoskeleton, sensing the environment, and motility. Deregulation of centrosome and cilium components leads to a wide range of diseases, some of which are incompatible with life. Centrosomes and cilia are thought to be very stable and can persist over long periods of time. However, these structures can disappear in certain developmental stages and diseases. Moreover, some centrosome and cilia components are quite dynamic. While a large body of knowledge has been produced regarding the biogenesis of these structures, little is known about how they are maintained. In this Review, we propose the existence of specific centrosome and cilia maintenance programs, which are regulated during development and homeostasis, and when deregulated can lead to disease., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2017. Published by The Company of Biologists Ltd.)

Centrioles are microtubule-based cylinders essential for the formation of centrosomes and cilia. A recent study provides a new cell-free assay that reconstitutes the initial structure formed during centriole assembly - the cartwheel - and proposes a new model for its formation and growth., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Centrioles and basal bodies (CBBs) organize centrosomes and cilia within eukaryotic cells. These organelles are composed of microtubules and hundreds of proteins performing multiple functions such as signaling, cytoskeleton remodeling, and cell motility. The CBB is present in all branches of the eukaryotic tree of life and, despite its ultrastructural and protein conservation, there is diversity in its function, occurrence (i.e., presence/absence), and modes of biogenesis across species. In this review, we provide an overview of the multiple pathways through which CBBs are formed in nature, with a special focus on the less studied, noncanonical ways. Despite the differences among each mechanism herein presented, we highlighted some of their common principles. These principles, governing different steps of biogenesis, ensure that CBBs may perform a multitude of functions in a huge diversity of organisms but yet retained their robustness in structure throughout evolution., (© 2017 Nabais et al.; Published by Cold Spring Harbor Laboratory Press.)

Barrett's esophagus is the major risk factor for esophageal adenocarcinoma. It has a low but non-neglectable risk, high surveillance costs and no reliable risk stratification markers. We sought to identify early biomarkers, predictive of Barrett's malignant progression, using a meta-analysis approach on gene expression data. This in silico strategy was followed by experimental validation in a cohort of patients with extended follow up from the Instituto Português de Oncologia de Lisboa de Francisco Gentil EPE (Portugal). Bioinformatics and systems biology approaches singled out two candidate predictive markers for Barrett's progression, CYR61 and TAZ. Although previously implicated in other malignancies and in epithelial-to-mesenchymal transition phenotypes, our experimental validation shows for the first time that CYR61 and TAZ have the potential to be predictive biomarkers for cancer progression. Experimental validation by reverse transcriptase quantitative PCR and immunohistochemistry confirmed the up-regulation of both genes in Barrett's samples associated with high-grade dysplasia/adenocarcinoma. In our cohort CYR61 and TAZ up-regulation ranged from one to ten years prior to progression to adenocarcinoma in Barrett's esophagus index samples. Finally, we found that CYR61 and TAZ over-expression is correlated with early focal signs of epithelial to mesenchymal transition. Our results highlight both CYR61 and TAZ genes as potential predictive biomarkers for stratification of the risk for development of adenocarcinoma and suggest a potential mechanistic route for Barrett's esophagus neoplastic progression., Competing Interests: Authors JC and JPL received funding from commercial company Ophiomics Precision Medicine, for this study. There are no patents, products in development or marketed products to declare. This does not alter our adherence to all the PLOS ONE policies on sharing data and materials.

The fruit fly, Drosophila melanogaster, is one of the most extensively studied organisms in biological research and has centrioles/basal bodies and cilia that can be modelled to investigate their functions in animals generally. Centrioles are nine-fold symmetrical microtubule-based cylindrical structures required to form centrosomes and also to nucleate the formation of cilia and flagella. When they function to template cilia, centrioles transition into basal bodies. The fruit fly has various types of basal bodies and cilia, which are needed for sensory neuron and sperm function. Genetics, cell biology and behaviour studies in the fruit fly have unveiled new basal body components and revealed different modes of assembly and functions of basal bodies that are conserved in many other organisms, including human, green algae and plasmodium. Here we describe the various basal bodies of Drosophila, what is known about their composition, structure and function.

Centrioles are essential for the assembly of both centrosomes and cilia. Centriole biogenesis occurs once and only once per cell cycle and is temporally coordinated with cell-cycle progression, ensuring the formation of the right number of centrioles at the right time. The formation of new daughter centrioles is guided by a pre-existing, mother centriole. The proximity between mother and daughter centrioles was proposed to restrict new centriole formation until they separate beyond a critical distance. Paradoxically, mother and daughter centrioles overcome this distance in early mitosis, at a time when triggers for centriole biogenesis Polo-like kinase 4 (PLK4) and its substrate STIL are abundant. Here we show that in mitosis, the mitotic kinase CDK1-CyclinB binds STIL and prevents formation of the PLK4-STIL complex and STIL phosphorylation by PLK4, thus inhibiting untimely onset of centriole biogenesis. After CDK1-CyclinB inactivation upon mitotic exit, PLK4 can bind and phosphorylate STIL in G1, allowing pro-centriole assembly in the subsequent S phase. Our work shows that complementary mechanisms, such as mother-daughter centriole proximity and CDK1-CyclinB interaction with centriolar components, ensure that centriole biogenesis occurs once and only once per cell cycle, raising parallels to the cell-cycle regulation of DNA replication and centromere formation., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Centriole elimination is an essential process that occurs in female meiosis of metazoa to reset centriole number in the zygote at fertilization. How centrioles are eliminated remains poorly understood. Here we visualize the entire elimination process live in starfish oocytes. Using specific fluorescent markers, we demonstrate that the two older, mother centrioles are selectively removed from the oocyte by extrusion into polar bodies. We show that this requires specific positioning of the second meiotic spindle, achieved by dynein-driven transport, and anchorage of the mother centriole to the plasma membrane via mother-specific appendages. In contrast, the single daughter centriole remaining in the egg is eliminated before the first embryonic cleavage. We demonstrate that these distinct elimination mechanisms are necessary because if mother centrioles are artificially retained, they cannot be inactivated, resulting in multipolar zygotic spindles. Thus, our findings reveal a dual mechanism to eliminate centrioles: mothers are physically removed, whereas daughters are eliminated in the cytoplasm, preparing the egg for fertilization., (© 2016 Borrego-Pinto et al.)

Centrioles and cilia are highly conserved eukaryotic organelles. Drosophila melanogaster is a powerful genetic and cell biology model organism, extensively used to discover underlying mechanisms of centrosome and cilia biogenesis and function. Defects in centrosomes and cilia reduce fertility and affect different sensory functions, such as proprioception, olfaction, and hearing. The fly possesses a large diversity of ciliary structures and assembly modes, such as motile, immotile, and intraflagellar transport (IFT)-independent or IFT-dependent assembly. Moreover, all the diverse ciliated cells harbor centrioles at the base of the cilia, called basal bodies, making the fly an attractive model to better understand the biology of this organelle. This chapter describes protocols to visualize centrosomes and cilia by fluorescence and electron microscopy.

Cilia are essential for cell signaling and sensory perception. In many cell types, a cytoskeletal structure called the ciliary rootlet links the cilium to the cell body. Previous studies indicated that rootlets support the long-term stability of some cilia. Here we report that Drosophila melanogaster Rootletin (Root), the sole orthologue of the mammalian paralogs Rootletin and C-Nap1, assembles into rootlets of diverse lengths among sensory neuron subtypes. Root mutant neurons lack rootlets and have dramatically impaired sensory function, resulting in behavior defects associated with mechanosensation and chemosensation. Root is required for cohesion of basal bodies, but the cilium structure appears normal in Root mutant neurons. We show, however, that normal rootlet assembly requires centrioles. The N terminus of Root contains a conserved domain and is essential for Root function in vivo. Ectopically expressed Root resides at the base of mother centrioles in spermatocytes and localizes asymmetrically to mother centrosomes in neuroblasts, both requiring Bld10, a basal body protein with varied functions., (© 2015 Chen et al.)