A network of spectrin and plectin surrounds the actin cuffs of apical tubulobulbar complexes in the rat

Abstract

Tubulobulbar complexes (TBCs) are actin-related endocytic structures that internalize intercellular junctions in the seminiferous epithelium. The structures consist of elongate tubular projections of the attached plasma membranes of two adjacent cells that project into Sertoli cells. This double membrane core is cuffed by a dentritic actin network and is capped at its end by a clathrin-coated pit. Here we explore the possibility that elements of the spectrin cytoskeleton are associated with clusters of tubulobulbar complexes that develop at adhesion junctions between late spermatids and Sertoli cells at the apex of the epithelium, and extend what is known about the distribution of plectin at the sites.Cryo-sections of perfusion-fixed testes and apical processes of Sertoli cells mechanically dissociated from perfusion-fixed testes were probed for spectrin, EPB41, and actin and analyzed using conventional fluorescence microscopy and confocal microscopy. Data sets from confocal microscopy were analyzed further in three-dimensional reconstructions using computer software. Additional apical Sertoli cell processes were probed for plectin and analyzed using conventional fluorescence microscopy. Antibodies generated against elements of the spectrin cytoskeleton react with material around and between the actin cuffs of tubulobulbar complexes, but appear excluded from the actin cuffs themselves. A similar staining pattern occurs with a probe for plectin. Immunoelectron microscopy confirmed the staining patterns observed by fluourescence microscopy. Based on our results, we suggest that a network of spectrin and plectin forms a scaffold around tubulobulbar complexes that may provide support for the actin network that cuffs each complex and also link adjacent complexes together.

Long non-coding RNA regulation of spermatogenesis and endosomal processes via the spectrin cytoskeleton inDrosophila

“…Herein we identify an uncharacterized lncRNA that interacts with α and β Spectrin. The spectrin cytoskeleton is modulated in axon development and degeneration 23,24 , myoblast fusion 25 , and spermatogenesis 12 through junctional protein complexes 26 , however, our work suggests lncRNAs can also interact with and regulate spectrin during these processes. We provide biochemical and genetic evidence that the targeted deletion mutation of the lncRNA CR45362 gene results in male sterility and alters endosomal related processes in the fat body of Drosophila melanogaster through α/β Spectrin disruption.…”

“…to migrate the maturing spermatocyte to the lumen of the seminiferous tubules and hold the spermatid until mature, all while maintaining the blood-testis barrier. 12 In Drosophila the male germ cells are totally encompasses and maintained by 2 Cyst cells, which are similar to human Sertoli cells 13 . The cyst cells provide molecular cues and provide protection until sperm maturate and are released into the seminal vesicle.…”

Functional importance of JMY expression by Sertoli cells in mediating mouse spermatogenesis

“…Considering the known actin-regulating activity of JMY, such we were prompted to determine if it interacts with these other actin cytoskeleton related proteins. Within this group, drebrin, actin-related protein 2/3 complex (Arp2/3), Anxa2, Cofilin, Gelsolin, spectrin have been reported to participate in regulating cell junction formation and function in Sertoli cells (Guttman et al, 2002; Lui et al, 2003; Lie et al, 2010; Li et al, 2011; Aristaeus de Asis et al, 2013; Chen et al, 2017; Chojnacka et al, 2017).…”

Long noncoding RNA regulation of spermatogenesis via the spectrin cytoskeleton in Drosophila

“…Herein, we identify an uncharacterized lncRNA CR45362 in Drosophila that interacts with α-Spectrin to stabilize spermatid nuclear bundles in the terminal epithelium region of basal testis. The spectrin cytoskeleton is modulated in axon development and degeneration ( Wang et al 2018 ; 2019 ), myoblast fusion ( Duan et al 2018 ), and spermatogenesis ( Aristaeus de Asis et al 2013 ) through junctional protein complexes ( Machnicka et al 2019 ), We found that CR45362 knockout results in male sterility, disrupted spermatid nuclear bundling and interaction between α-Spectrin and actin cap. Our results indicate a novel mechanism by which the spectrin cytoskeleton can be contextually regulated through lncRNAs.…”

“…During mammalian sperm maturation, Sertoli cells surround developing sperm and undergo intricate endocytic exchanges of intercellular junctions (ectoplasmic specialization) and tubulobulbar complexes (actin-spectrin-clathrin structures) to facilitate the movement of spermatocytes towards the lumen of the seminiferous tubules and hold the spermatid until mature, while maintaining the blood-testis barrier ( Aristaeus de Asis et al 2013 ). In Drosophila the male germ cells are encompassed and maintained by 2 Cyst cells, which are similar to human Sertoli cells ( Zoller and Schulz 2012 ).…”