An intracellular trafficking pathway in the seminiferous epithelium regulating spermatogenesis: a biochemical and molecular perspective

During spermatogenesis in adult rat testes, fully developed spermatids (i.e. spermatozoa) at the luminal edge of the seminiferous epithelium undergo “spermiation” at stage VIII of the seminiferous epithelial cycle. This is manifested by the disruption of the apical ectoplasmic specialization (apical ES) so that spermatozoa can enter the tubule lumen and to complete their maturation in the epididymis. At the same time, the blood–testis barrier (BTB) located near the basement membrane undergoes extensive restructuring to allow transit of preleptotene spermatocytes so that post-meiotic germ cells complete their development behind the BTB. While spermiation and BTB restructuring take place concurrently at opposite ends of the Sertoli cell epithelium, the biochemical mechanism(s) by which they are coordinated were not known until recently. Studies have shown that fragments of laminin chains are generated from the laminin/integrin protein complex at the apical ES via the action of MMP-2 (matrix metalloprotease-2) at spermiation. These peptides serve as the local autocrine factors to destabilize the BTB. These laminin peptides also exert their effects on hemidesmosome which, in turn, further potentiates BTB restructuring. Thus, a novel apical ES-BTB-hemidesmosome regulatory loop is operating in the seminiferous epithelium to coordinate these two crucial cellular events of spermatogenesis. This functional loop is further assisted by the Par3/Par6-based polarity protein complex in coordination with cytokines and testosterone at the BTB. Herein, we provide a critical review based on the latest findings in the field regarding the regulation of these cellular events. These recent findings also open up a new window for investigators studying blood–tissue barriers.     

Wnt and planar cell polarity signaling in cystic renal disease

Cystic kidney diseases can cause end stage renal disease, affecting millions of individuals worldwide. They may arise early or later in life, are characterized by a spectrum of symptoms and can be caused by diverse genetic defects. The primary cilium, a microtubule-based organelle that can serve as a signaling antenna, has been demonstrated to have a significant role in ensuring correct kidney development and function. In the kidney, one of the signaling pathways that requires the cilium for normal development is Wnt signaling. In this review, the roles of primary cilia in relation to canonical and non-canonical Wnt/PCP signaling in cystic renal disease are described. The evidence of the associations between cilia, Wnt signaling and cystic renal disease is discussed and the significance of planar cell polarity-related mechanisms in cystic kidney disease is presented. Although defective Wnt signaling is not the only cause of renal disease, research is increasingly highlighting its importance, encouraging the development of Wnt-associated diagnostic and prognostic tools for cystic renal disease.     

Wnt and planar cell polarity signaling in cystic renal disease

Cystic kidney diseases can cause end stage renal disease, affecting millions of individuals worldwide. They may arise early or later in life, are characterized by a spectrum of symptoms and can be caused by diverse genetic defects. The primary cilium, a microtubule-based organelle that can serve as a signaling antenna, has been demonstrated to have a significant role in ensuring correct kidney development and function. In the kidney, one of the signaling pathways that requires the cilium for normal development is Wnt signaling. In this review, the roles of primary cilia in relation to canonical and non-canonical Wnt/PCP signaling in cystic renal disease are described. The evidence of the associations between cilia, Wnt signaling and cystic renal disease is discussed and the significance of planar cell polarity-related mechanisms in cystic kidney disease is presented. Although defective Wnt signaling is not the only cause of renal disease, research is increasingly highlighting its importance, encouraging the development of Wnt-associated diagnostic and prognostic tools for cystic renal disease.     

Control of planar cell polarity by interaction of DWnt4 and four-jointed

The Drosophila eye and the wing display specific planar cell polarity. Although Frizzled (Fz) signaling has been implicated in the establishment of ommatidial and wing hair polarity, evidence for the Wnt gene function has been limited. Here we examined the function of a Drosophila homolog of Wnt4 (DWnt4) in the control of planar polarity. We show that DWnt4 mRNA and protein are preferentially expressed in the ventral region of eye disc. DWnt4 mutant eyes show polarity reversals mostly in the ventral domain, consistent with the ventral expression of DWnt4. Ectopic expression of DWnt4 in the dorsoventral (DV) polar margins is insufficient to induce ommatidial polarity but becomes inductive when coexpressed with Four-jointed (Fj). Similarly, DWnt4 and Fj result in synergistic induction of hair polarity toward the source of expression in the wing. Consistent with genetic interaction, we provide evidence for direct interaction of DWnt4 and Fj transmembrane protein. The extracellular domain of Fj is required for direct binding to DWnt4 and for the induction of hair polarity. In contrast to the synergy between DWnt4 and Fj, DWnt4 antagonizes the polarizing effect of Fz. Our results suggest that DWnt4 is involved in ommatidial polarity signaling in the ventral region of the eye and its function is mediated by interacting with Fj.     

Roles of planar cell polarity pathways in the development of neutral tube defects

Neural tube defects (NTDs) are the second most common birth defect in humans. Despite many advances in the understanding of NTDs and the identification of many genes related to NTDs, the fundamental etiology for the majority of cases of NTDs remains unclear. Planar cell polarity (PCP) signaling pathway, which is important for polarized cell movement (such as cell migration) and organ morphogenesis through the activation of cytoskeletal pathways, has been shown to play multiple roles during neural tube closure. The disrupted function of PCP pathway is connected with some NTDs. Here, we summarize our current understanding of how PCP factors affect the pathogenesis of NTDs.     

Insight into planar cell polarity

Planar cell polarity or PCP refers to a uniform cellular organization within the plan, typically orthogonal to the apico-basal polarity axis. As such, PCP provides directional cues that control and coordinate the integration of cells in tissues to build a living organism. Although dysfunctions of this fundamental cellular process have been convincingly linked to the etiology of various pathologies such as cancer and developmental defects, the molecular mechanisms governing its establishment and maintenance remain poorly understood. Here, we review some aspects of invertebrate and vertebrate PCPs, highlighting similarities and differences, and discuss the prevalence of the non-canonical Wnt signaling as a central PCP pathway, as well as recent findings on the importance of cell contractility and cilia as promising avenues of investigation.     

Sertoli cell Dicer is essential for spermatogenesis in mice

Spermatogenesis requires intact, fully competent Sertoli cells. Here, we investigate the functions of Dicer, an RNaseIII endonuclease required for microRNA and small interfering RNA biogenesis, in mouse Sertoli cell function. We show that selective ablation of Dicer in Sertoli cells leads to infertility due to complete absence of spermatozoa and progressive testicular degeneration. The first morphological alterations appear already at postnatal day 5 and correlate with a severe impairment of the prepubertal spermatogenic wave, due to defective Sertoli cell maturation and incapacity to properly support meiosis and spermiogenesis. Importantly, we find several key genes known to be essential for Sertoli cell function to be significantly down-regulated in neonatal testes lacking Dicer in Sertoli cells. Overall, our results reveal novel essential roles played by the Dicer-dependent pathway in mammalian reproductive function, and thus pave the way for new insights into human infertility.     

Drosophila ATP6AP2/VhaPRR functions both as a novel planar cell polarity core protein and a regulator of endosomal trafficking

Planar cell polarity (PCP) controls the orientation of cells within tissues and the polarized outgrowth of cellular appendages. So far, six PCP core proteins including the transmembrane proteins Frizzled (Fz), Strabismus (Stbm) and Flamingo (Fmi) have been identified. These proteins form asymmetric PCP domains at apical junctions of epithelial cells. Here, we demonstrate that VhaPRR, an accessory subunit of the proton pump V‐ATPase, directly interacts with the protocadherin Fmi through its extracellular domain. It also shows a striking co‐localization with PCP proteins during all pupal wing stages in Drosophila. This localization depends on intact PCP domains. Reversely, VhaPRR is required for stable PCP domains, identifying it as a novel PCP core protein. VhaPRR performs an additional role in vesicular acidification as well as endolysosomal sorting and degradation. Membrane proteins, such as E‐Cadherin and the Notch receptor, accumulate at the surface and in intracellular vesicles of cells mutant for VhaPRR. This trafficking defect is shared by other V‐ATPase subunits. By contrast, the V‐ATPase does not seem to have a direct role in PCP regulation. Together, our results suggest two roles for VhaPRR, one for PCP and another in endosomal trafficking. This dual function establishes VhaPRR as a key factor in epithelial morphogenesis.     

Left‐right asymmetry in the chick embryo requires core planar cell polarity protein Vangl2

Consistent left‐right patterning is a fascinating and biomedically important problem. In the chick embryo, it is not known how cells determine their position (left or right) relative to the primitive streak, which is required for subsequent asymmetric gene expression cascades. We show that the subcellular localization of Vangl2, a core planar cell polarity (PCP) protein, is consistently polarized, giving cells in the blastoderm a vector pointing toward the primitive streak. Moreover, morpholino‐mediated loss‐of‐function of Vangl2 by electroporation into chicks at very early stages randomizes the normally left‐sided expression of Sonic hedgehog. Strikingly, Vangl2 morpholinos also induce a desynchronization of asymmetric gene expression within the left and right domains of Hensen's node. These data reveal the existence of polarized planar cell polarity protein localization in gastrulating chick and demonstrate that the PCP pathway is functionally required for normal asymmetry in the chick upstream of Sonic hedgehog. These data suggest a new and widely applicable class of models for the spread and coordination of left‐right patterning information in the embryonic blastoderm. genesis 47:719–728, 2009. © 2009 Wiley‐Liss, Inc.     

Hormonal induction of all stages of spermatogenesis in germ-somatic cell coculture from immature Japanese eel testis

In cultivated male eel, spermatogonia are the only germ cells present in testis. Our previous studies using an organ culture system have shown that gonadotropin and 11-ketotestosterone (11-KT, a potent androgen in teleost fishes) can induce all stages of spermatogenesis in vitro. for detailed investigation of the control mechanisms of spermatogenesis, especially of the interaction between germ cells and testicular somatic cells during 11-KT-induced spermatogenesis in vitro, we have established a new culture system in which germ cells and somatic cells are cocultured after they are aggregated into pellets by centrifugation. Germ cells (spermatogonia) and somatic cells (mainly Sertoli cells) were isolated from immature eel testis. Coculture of the isolated germ cells and somatic cells without forming aggregation did not induce spermatogenesis, even in the presence of 11-KT. In contrast, when isolated germ cells and somatic cells were formed into pellets by centrifugation and were then cultured with 11-KT for 30 days, the entire process of spermatogenesis from premitotic spermatogonia to spermatozoa was induced. However, in the absence of 11-KT in the culture medium spermatogenesis was not induced, even when germ cell and somatic cells were aggregated. These results demonstrate that physical contact of germ cells to Sertoli cells is required for inducing spermatogenesis in response to 11-KT.