Disruption of sperm release from insect testes by cytochalasin and ?-actin mRNA mediated interference

Abstract

Release of sperm bundles from moth testes is controlled by the local circadian oscillator. The mechanism which restricts migration of sperm bundles to a few hours each day is not understood. We demonstrate that a daily cycle of sperm release is initiated by the migration of folded apyrene sperm bundles through a cellular barrier at the testis base. These bundles have conspicuous concentrations of actin filaments at their proximal end. Inhibition of actin polymerization by cytochalasin at aspecific time of day inhibited sperm release from the testis. Likewise, application of double-stranded actin RNA specifically inhibited sperm release. This RNA-mediated interference (RNAi) lowered the pool of actin mRNA in tissues involved in sperm release. The decline in mRNA levels resulted in the selective depletion of F-actin from the tip of apyrene sperm bundles, suggesting that this actin may be involved in the initiation of sperm release. Combined results of RNAi experiments at physiological, cellular and molecular levels identified unique cells that are critically involved in the mechanism of sperm release.

F-actin-based extensions of the head cyst cell adhere to the maturing spermatids to maintain them in a tight bundle and prevent their premature release in Drosophila testis

“…The conjugated NB and IC disruptions reflected disorganizations of the spermatid bundles inside the cysts and the rapid manifestation of the defect further suggested that the association between the head cyst cell and the spermatids are dynamic. F-actin disruption in isolated testis of adult moths was shown to block sperm release and spermatid bundle disorganization [44] and studies in mammalian testis showed that the F-actin stability in the apical ES is essential to maintaining association between developing spermatids and Sertoli cells [12]. …”

Effects of larval diapause and the juvenile hormone analog, fenoxycarb, on testis development and spermatogenesis in the wax moth, Galleria mellonella (Lepidoptera: Pyralidae)

“…This also occurs in B. mori (Katsuno, 1977a), Lymantria dispar (Giebultowicz & Joy, 1992;Giebultowicz et al, 1997) and S. littoralis (Bebas et al, 2001); however, its role remains unknown. Because sperm transfer to the UVD is a complex process involving interactions between the cyst cells of sperm bundles and the epithelial cells of testes (forming the cellular membrane between the TF and UVD), it is suggested that apyrene bundles somehow initiate this process for eupyrene bundles (Gvakharia et al, 2003). This explanation is strongly supported by the features of the transfer of sperm from the testis to the UVD in adults of E. kuehniella (Riemann et al, 1974), B. mori (Katsuno, 1977b, S. littoralis (Bebas et al, 2001) and S. litura (Seth et al, 2002), which is described as a repeatedly occurring event, initiated daily by apyrene sperm transfer.…”

“…entiation into two types, leading to the formation of nucleated (eupyrene) and un-nucleated (apyrene) spermatozoa (Mancini & Dolder, 2004;Friedländer et al, 2005;Dallai, 2014;Dallai et al, 2016). During release from the testis into the upper vasa deferentia (UVD), both types of spermatozoa are liberated from cyst cells that remain in the testis and then undergo programmed cell death followed by fragmentation (Giebultowicz et al, 1997;Gvakharia et al, 2003). In the UVD, apyrene spermatozoa in each bundle are associated for a short time after release and then disperse, whereas eupyrene spermatozoa remain in tight bundles during transfer in the reproductive ducts of males (Riemann & Giebultowicz, 1992;Friedländer et al, 2001).…”

Functional analysis of the allatostatin-A type gene in the cricket Gryllus bimaculatus and the armyworm Spodoptera frugiperda

“…There was some remaining gene activity, which means that the silencing, which is a posttranscriptional process in animals , was incomplete. This was also observed after injection of dsRNA into adult flies against Toll pathway components (Goto et al, 2003) or by tissue incubation or dsRNA injection of moths, ticks and honeybees (Gvakharia et al, 2003;Karim et al, 2004;Farooqui et al, 2004), where gene expression was measured either by semiquantitative RT-PCR or by Western blot of the target proteins. In contrast, in adult honeybees the vitellogenin gene (Amdam et al, 2003) and in larval Spodoptera litura the midgut aminopeptidase N gene (Rajagopal et al, 2002) could be almost totally silenced by abdominal injections of dsRNA.…”