Biochem. J. (2011) 435, 553–56 2 (Printed in Great Britain) doi:10.1042 /BJ20102121 553 REVIEW ARTICLE Regulation of spermiogenesis, spermiation and blood–testis barrier dynamics: novel insights from studies on Eps8 and Arp3 C. Yan CHENG 1 and Dolores D. MRUK The Mary M. Wohlford Laboratory for Male Contraceptive Research, Center for Biomedical Research, Population Council, 1230 York Avenue, New York, NY 10065, U.S.A. Spermiogenesis in the mammalian testis is the most critical post- meiot ic dev elopmental eve nt occur ring durin g spermatogen esis in which haploid spermatids undergo extensive cellular, molecular and morphological changes to form spermatozoa. Spermatozoa are the n rel eas ed from the semini fer ous epi the liu m at spe rmi ati on. At the same time, the BTB (bl ood–te stis bar rie r) und erg oes restructuri ng to facil itate the trans it of prele ptoten e spermatoc ytes from the basal to the apical compartment. Thus meiotic divisions take place behind the BTB in the apical compartment to form spermatids. These germ cells enter spermiogenesis to transform into elongating spermatids and then into spermatozoa to replace those tha t wer e rel eas ed in the pre vio us cyc le. Howe ve r, the mol e- cular regulato rs that control spermi ogenes is, in partic ular the dynami c cha nge s tha t occ ur at the Ser tol i cel l–s per mat id int erf ace and at the BTB, are not entirely known. This is largely due to the lack of suitable animal models which can be used to study these events. During the course of our investigation to develop adjudin [1-(2,4-dichlorobenzyl)- 1H-indazole-3-carbohydrazide] as a potent ia l male cont race pt iv e, this dr ug wa s shown to ‘acce lerat e’ spermiation by induc ing the release of prema ture sperma tids fro m the epi the lium. Usi ng thi s mod el, we ha ve ident ified severa l molec ules that are crucial in regul ating the actin filament network and the unique adhesion protein complex at the Sertoli cell–spermatid interface known as the apical ES (ectoplasmic specialization). In the present review, we critically eval ua te these and othe r findi ngs in the litera ture as they relate to the restricted temporal and spatial expression of two actin regulatory proteins, namely Eps8 (epidermal growth factor receptor pathway substrate 8) and Arp3 (actin-related protein 3), which regulate these events. Key words: adjudin, blood–testis barrier, intermediate filament, semin ifero us epithelial cycle, spermatogenesis, tubulobulbar complex. INTRODUCTION Most cha nge s in cell mor pho logy , pla sticit y and mov eme nt resulting from cues received from the environment, growth and development, stress, cytokines, toxicants or during pathogenesis are re gulate d by the ac ti n- , inte rmediate fil ament- and/ or tubulin-based cytoskeletons [ 1–6]. This applies to virtually all epithelial cells, including those in the seminiferous epithelium of the mammalian testis. Interestingly, the actin network in the seminiferous epithelium, which is composed of only Sertoli cells and germ cells which are at different stages of development (i.e. spermatogon ial stem cells,spermatogo nia, prima ry and secon dary spermatocy tes, spermatids and spermatozo a) (Figu re 1), is not abl y different from actin networks found in other epithelia in several way s. Fir st, actin fila men t bundl es found in Ser tol i cel ls at the ES {ectop lasmi c speci aliza tion; a testis -speci fic atypi cal AJ (adherens junction) found at the Sertoli cell–spermatid and Sertoli–Sertoli cell interface known as the apical and basal ES respectively[7–9]} (Figures 1 and 2) are non-contractile [9,10], even thoug h motor protein s (e.g. myosin VIIa) are prese nt at the ES [10,11]. Sec ondly , act in fila men t bundl es are tig htl y pac ked , arr ang ed par all el to theSertoli cel l pla smamembr ane , and sandwiched in between apposing cell membranes of Sertoli cell– spermatid or Sertol i–Sert oli cell and cister nae of endop lasmi c reticulum(Figures 1 and 2). Although other studies have shown that the ES is one of the strongest adhesive junctions [12,13], it sti ll is sub jec ted to ext ens iv e res tructu ring. Thi s thu s fa cil ita tes the trans it of sperma tids acros s the epith elium durin g spermiogen esis, as well as the transit of preleptotene spermatocytes across the BTB (blood–te sti s bar rie r) at sta ge VIII of the seminiferous epith elial cycl e of sperma togen esis [14,15]. Third ly , altho ugh Sertoli cells cultu red in vitr o are highly mot ile and cap able of traversing the membranes of transwell (i.e. bicameral) units simil ar to metast atic cancer cells [16,17], Ser tol i cel ls are in fac t not mot ile in viv o. Ins tead, they are static , ‘nurs e-lik e’ cells needed for germ cell development with each Sertoli cell ‘engulfing’ approx. 30–50 developing germ cells, but they do alter their cell shape to acommodate morphological changes ofspermatids durin g spermiogene sis. Moreo ver , Serto li cells are the only struct ural and ‘scaf foldi ng’ cells in the seminifero us epithelium that confer BTB function via co-existing TJs (tight junctions), basal ES, desmosomes and gap junctions located near the basement membrane in the seminiferous epithelium, since microvessels in the interstitial space between tubules contribute relatively little to the BTB. On a final note, all of these junctions link to either the actin, the intermediate filament or the tubulin network, and they are inter conne cted struct urall y and funct ionall y. In the present review, we critically discuss results from recent studies relating to two actin regulatory proteins: Eps8 (epidermal growth factor receptor pathway substrate 8) ( Figure 3) and Arp3 Abbreviations used: adjudin, 1-(2,4-dichlorobenzyl)- 1H-indazole-3-carbohydrazide; AJ, adherens junction; Arp, actin-related protein; ARPC, Arp2/3 complex subunit; BTB, blood–testis barrier; Cdc42, cell division cycle 42; Eps8, epidermal growth factor receptor pathway substrate 8; ES, ectoplasmic specialization; F-actin, filamentous actin; PAR, partitioning defective protein; N-WASP , neuronal WASP; SCAR/WA VE, suppressor of cAMP receptor/WASP family verprolin homologous; TJ, tight junction; WASP, Wiskott–Aldrich syndrome protein. 1 T o whom corresponden ce should be addressed (email Y -Cheng@popcbr .rockefeller .edu). c The Authors Journal compilation c 2011 Biochemical Society www.biochemj.org B i o c h e m i c a l J o u r n a l
