Centrosome positioning in interphase cells

The position of the centrosome is actively maintained at the cell center, but the mechanisms of the centering force remain largely unknown. It is known that centrosome positioning requires a radial array of cytoplasmic microtubules (MTs) that can exert pushing or pulling forces involving MT dynamics and the activity of cortical MT motors. It has also been suggested that actomyosin can play a direct or indirect role in this process. To examine the centering mechanisms, we introduced an imbalance of forces acting on the centrosome by local application of an inhibitor of MT assembly (nocodazole), and studied the resulting centrosome displacement. Using this approach in combination with microinjection of function-blocking probes, we found that a MT-dependent dynein pulling force plays a key role in the positioning of the centrosome at the cell center, and that other forces applied to the centrosomal MTs, including actomyosin contractility, can contribute to this process.     

EB1 is required for primary cilia assembly in fibroblasts

EB1 is a small microtubule (MT)-binding protein that associates preferentially with MT plus ends and plays a role in regulating MT dynamics. EB1 also targets other MT-associated proteins to the plus end and thereby regulates interactions of MTs with the cell cortex, mitotic kinetochores, and different cellular organelles [1, 2]. EB1 also localizes to centrosomes and is required for centrosomal MT anchoring and organization of the MT network [3, 4]. We previously showed that EB1 localizes to the flagellar tip and proximal region of the basal body in Chlamydomonas[5], but the function of EB1 in the cilium/flagellum is unknown. We depleted EB1 from NIH3T3 fibroblasts by using siRNA and found that EB1 depletion causes a approximately 50% reduction in the efficiency of primary cilia assembly in serum-starved cells. Expression of dominant-negative EB1 also inhibited cilia formation, and expression of mutant dominant-negative EB1 constructs suggested that binding of EB1 to p150(Glued) is important for cilia assembly. Finally, expression of a C-terminal fragment of the centrosomal protein CAP350, which removes EB1 from the centrosome but not MT plus ends [6], also inhibited ciliogenesis. We conclude that localization of EB1 at the centriole/basal body is required for primary cilia assembly in fibroblasts.     

Trichoplein and Aurora A block aberrant primary cilia assembly in proliferating cells

The primary cilium is an antenna-like organelle that modulates differentiation, sensory functions, and signal transduction. After cilia are disassembled at the G0/G1 transition, formation of cilia is strictly inhibited in proliferating cells. However, the mechanisms of this inhibition are unknown. In this paper, we show that trichoplein disappeared from the basal body in quiescent cells, whereas it localized to mother and daughter centrioles in proliferating cells. Exogenous expression of trichoplein inhibited primary cilia assembly in serum-starved cells, whereas ribonucleic acid interference-mediated depletion induced primary cilia assembly upon cultivation with serum. Trichoplein controlled Aurora A (AurA) activation at the centrioles predominantly in G1 phase. In vitro analyses confirmed that trichoplein bound and activated AurA directly. Using trichoplein mutants, we demonstrate that the suppression of primary cilia assembly by trichoplein required its ability not only to localize to centrioles but also to bind and activate AurA. Trichoplein or AurA knockdown also induced G0/G1 arrest, but this phenotype was reversed when cilia formation was prevented by simultaneous knockdown of IFT-20. These data suggest that the trichoplein-AurA pathway is required for G1 progression through a key role in the continuous suppression of primary cilia assembly.     

Qilin is essential for cilia assembly and normal kidney development in zebrafish

Defects in the cilium, a once thought vestigial organelle, have recently been implicated in many human diseases, including a number of cystic kidney diseases such as polycystic kidney disease (PKD), Bardet Bieldl Syndrome, and Meckel-Gruber Syndrome. In a forward genetic screen, qilin was identified as a novel gene important in the pathogenesis of kidney cysts in zebrafish. In this paper we characterized qilin(hi3959A) mutant's phenotypes in detail, investigated cilia formation in this mutant and performed structural and functional analysis of the Qilin protein. Results reveal Qilin's essential role in cilia assembly and maintenance in multiple organs, including the kidney, the lateral line organ, and the outer segment of the photoreceptor cell. In addition, rescue experiments suggest that defective pronephric cilia correlate with the formation of kidney cysts in qilin(hi3959A) mutants. Further, genetic analysis suggests that qilin interacts with multiple intraflagellar transport (IFT) complex B genes, which is supported by the striking phenotypic similarities between qilin(hi3959A) and IFT complex B mutants. Finally, through deletion analysis we provide evidence that the well-conserved N-terminus and the coiled-coil domain of Qilin are both essential and sufficient for its function. Taken all the observations together, we propose that Qilin acts in a similar role as IFT complex B proteins in cilia assembly, maintenance and kidney development in zebrafish.     

Centrosome positioning in polarized cells: Common themes and variations

The centrosome position is tightly regulated during the cell cycle and during differentiated cellular functions. Because centrosome organizes the microtubule network to coordinate both intracellular organization and cell signaling, centrosome positioning is crucial to determine either the axis of cell division, the direction of cell migration or the polarized immune response of lymphocytes. Since alteration of centrosome positioning seems to promote cell transformation and tumor spreading, the molecular mechanisms controlling centrosome movement in response to extracellular and intracellular cues are under intense investigation. Evolutionary conserved pathways involving polarity proteins and cytoskeletal rearrangements are emerging as common regulators of centrosome positioning in a wide variety of cellular contexts.     

Kinesin motors and primary cilia

Cilia and flagella play important roles in human health by contributing to cellular motility as well as sensing and responding to environmental cues. Defects in ciliary assembly and/or function can lead to a range of human diseases, collectively known as the ciliopathies, including polycystic kidney, liver and pancreatic diseases, sterility, obesity, situs inversus, hydrocephalus and retinal degeneration. A basic understanding of how cilia form and function is essential for deciphering ciliopathies and generating therapeutic treatments. The cilium is a unique compartment that contains a distinct complement of protein and lipid. However, the molecular mechanisms by which soluble and membrane protein components are targeted to and trafficked into the cilium are not well understood. Cilia are generated and maintained by IFT (intraflagellar transport) in which IFT cargoes are transported along axonemal microtubules by kinesin and dynein motors. A variety of genetic, biochemical and cell biological approaches has established the heterotrimeric kinesin-2 motor as the 'core' IFT motor, whereas other members of the kinesin-2, kinesin-3 and kinesin-4 families function as 'accessory' motors for the transport of specific cargoes in diverse cell types. Motors of the kinesin-9 and kinesin-13 families play a non-IFT role in regulating ciliary beating or axonemal length, respectively. Entry of kinesin motors and their cargoes into the ciliary compartment requires components of the nuclear import machinery, specifically importin-β2 (transportin-1) and Ran-GTP (Ran bound to GTP), suggesting that similar mechanisms may regulate entry into the nuclear and ciliary compartments.     

Autophagy and regulation of cilia function and assembly

Motile and primary cilia (PC) are microtubule-based structures located at the cell surface of many cell types. Cilia govern cellular functions ranging from motility to integration of mechanical and chemical signaling from the environment. Recent studies highlight the interplay between cilia and autophagy, a conserved cellular process responsible for intracellular degradation. Signaling from the PC recruits the autophagic machinery to trigger autophagosome formation. Conversely, autophagy regulates ciliogenesis by controlling the levels of ciliary proteins. The cross talk between autophagy and ciliated structures is a novel aspect of cell biology with major implications in development, physiology and human pathologies related to defects in cilium function.     

Pleiotropic signaling pathways orchestrate yeast development

Developmental phenotypes in Saccharomyces cerevisiae and related yeasts include responses such as filamentous growth, sporulation, and the formation of biofilms and complex colonies. These developmental phenotypes are regulated by evolutionarily conserved, nutrient-responsive signaling networks. The signaling mechanisms that control development in yeast are highly pleiotropic--all the known pathways contribute to the regulation of multiple developmental outcomes. This degree of pleiotropy implies that perturbations of these signaling pathways, whether genetic, biochemical, or environmentally induced, can manifest in multiple (and sometimes unexpected) ways. We summarize the current state of knowledge of developmental pleiotropy in yeast and discuss its implications for understanding functional relationships.     

The radial organization of neuronal primary cilia is acutely disrupted by seizure and ischemic brain injury

Background

Neuronal primary cilia are sensory organelles that are critically involved in the proper growth, development, and function of the central nervous system (CNS). Recent work also suggests that they signal in the context of CNS injury, and that abnormal ciliary signaling may be implicated in neurological diseases.

Methods

We quantified the distribution of neuronal primary cilia alignment throughout the normal adult mouse brain by immunohistochemical staining for the primary cilia marker adenylyl cyclase III (ACIII) and measuring the angles of primary cilia with respect to global and local coordinate planes. We then introduced two different models of acute brain insult—temporal lobe seizure and cerebral ischemia, and re-examined neuronal primary cilia distribution, as well as ciliary lengths and the proportion of neurons harboring cilia.

Results

Under basal conditions, cortical cilia align themselves radially with respect to the cortical surface, while cilia in the dentate gyrus align themselves radially with respect to the granule cell layer. Cilia of neurons in the striatum and thalamus, by contrast, exhibit a wide distribution of ciliary arrangements. In both cases of acute brain insult, primary cilia alignment was significantly disrupted in a region-specific manner, with areas affected by the insult preferentially disrupted. Further, the two models promoted differential effects on ciliary lengths, while only the ischemia model decreased the proportion of ciliated cells.

Conclusions

These findings provide evidence for the regional anatomical organization of neuronal primary cilia in the adult brain and suggest that various brain insults may disrupt this organization.

     

Centrosome asymmetry and inheritance during animal development

The centrosome is a subcellular organelle that is responsible for the majority of microtubule organization. Through this ability, the centrosome is involved in cell division, migration, and polarization. Recent studies have revealed intriguing asymmetries between mother and daughter centrioles as well as between mother and daughter centrosomes, and the involvement of such asymmetries in multiple cellular and developmental processes. This review aims to summarize recent discoveries on such asymmetries in centrioles/centrosomes and the potential implication of their inheritance patterns during cell division and development.     

The role of primary cilia in the development and disease of the retina

The normal development and function of photoreceptors is essential for eye health and visual acuity in vertebrates. Mutations in genes encoding proteins involved in photoreceptor development and function are associated with a suite of inherited retinal dystrophies, often as part of complex multi-organ syndromic conditions. In this review, we focus on the role of the photoreceptor outer segment, a highly modified and specialized primary cilium, in retinal health and disease. We discuss the many defects in the structure and function of the photoreceptor primary cilium that can cause a class of inherited conditions known as ciliopathies, often characterized by retinal dystrophy and degeneration, and highlight the recent insights into disease mechanisms.     

The role of primary cilia in the development and disease of the retina

The normal development and function of photoreceptors is essential for eye health and visual acuity in vertebrates. Mutations in genes encoding proteins involved in photoreceptor development and function are associated with a suite of inherited retinal dystrophies, often as part of complex multi-organ syndromic conditions. In this review, we focus on the role of the photoreceptor outer segment, a highly modified and specialized primary cilium, in retinal health and disease. We discuss the many defects in the structure and function of the photoreceptor primary cilium that can cause a class of inherited conditions known as ciliopathies, often characterized by retinal dystrophy and degeneration, and highlight the recent insights into disease mechanisms.     

Primary cilia and forebrain development

With a microtubule-based axoneme supporting its plasma membrane-ensheathed projection from the basal body of almost all cell types in the human body, and present in only one copy per cell, the primary cilium can be considered an organelle sui generis. Although it was first observed and recorded in histological studies from the late 19th century, the tiny structure was essentially forgotten for many decades. In the past ten years, however, scientists have turned their eyes once again upon primary cilia and realized that they are very important for the development of almost all organs in the mammalian body, especially those dependent upon the signaling from members Hedgehog family, such as Indian and Sonic hedgehog. In this review, we outline the roles that primary cilia play in forebrain development, not just in the crucial transduction of Sonic hedgehog signaling, but also new results showing that cilia are important for cell cycle progression in proliferating neural precursors. We will focus upon cerebral cortex development but will also discuss the importance of cilia for the embryonic hippocampus, olfactory bulb, and diencephalon.