The dynamics and regulation of chromatin remodeling during spermiogenesis.

In most animals, the extreme compaction of sperm DNA is achieved after the massive replacement of histones with sperm nuclear basic proteins (SNBPs), such as protamines. In some species, the ultracompact sperm chromatin is stabilized by a network of disulfide bonds connecting cysteine residues present in SNBPs. Studies in mammals have established that the reduction of these disulfide crosslinks at fertilization is required for sperm nuclear decondensation and the formation of the male pronucleus. Here, we show that the Drosophila maternal thioredoxin Deadhead (DHD) is specifically required to unlock sperm chromatin at fertilization. In dhd mutant eggs, the sperm nucleus fails to decondense and the replacement of SNBPs with maternally-provided histones is severely delayed, thus preventing the participation of paternal chromosomes in embryo development. We demonstrate that DHD localizes to the sperm nucleus to reduce its disulfide targets and is then rapidly degraded after fertilization.

/pdf/unlocking-sperm-chromatin-at-fertilization-requires-a-4pj602mp7i.pdf

Unlocking sperm chromatin at fertilization requires a dedicated egg thioredoxin in Drosophila.

Tandemly-repeated DNAs, or satellites, are enriched in heterochromatic regions of eukaryotic genomes and contribute to nuclear structure and function. Some satellites are transcribed, but we lack direct evidence that specific satellite RNAs are required for normal organismal functions. Here, we show satellite RNAs derived from AAGAG tandem repeats are transcribed in many cells throughout Drosophila melanogaster development, enriched in neurons and testes, often localized within heterochromatic regions, and important for viability. Strikingly, we find AAGAG transcripts are necessary for male fertility, and that AAGAG RNA depletion results in defective histone-protamine exchange, sperm maturation and chromatin organization. Since these events happen late in spermatogenesis when the transcripts are not detected, we speculate that AAGAG RNA in primary spermatocytes 'primes' post-meiosis steps for sperm maturation. In addition to demonstrating essential functions for AAGAG RNAs, comparisons between closely related Drosophila species suggest that satellites and their transcription evolve quickly to generate new functions.

/pdf/rna-from-a-simple-tandem-repeat-is-required-for-sperm-iucm3g37ic.pdf

RNA from a simple-tandem repeat is required for sperm maturation and male fertility in Drosophila melanogaster.

In most animals, the bulk of sperm DNA is packaged with sperm nuclear basic proteins (SNBPs), a diverse group of highly basic chromosomal proteins notably comprising mammalian protamines. The replacement of histones with SNBPs during spermiogenesis allows sperm DNA to reach an extreme level of compaction, but little is known about how SNBPs actually function in vivo . Mst77F is a Drosophila SNBP with unique DNA condensation properties in vitro , but its role during spermiogenesis remains unclear. Here, we show that Mst77F is required for the compaction of sperm DNA and the production of mature sperm, through its cooperation with protamine-like proteins Mst35Ba/b. We demonstrate that Mst77F is incorporated in spermatid chromatin as a precursor protein, which is subsequently processed through the proteolysis of its N-terminus. The cleavage of Mst77F is very similar to the processing of protamine P2 during human spermiogenesis and notably leaves the cysteine residues in the mature protein intact, suggesting that they participate in the formation of disulfide cross-links. Despite the rapid evolution of SNBPs, sperm chromatin condensation thus involves remarkably convergent mechanisms in distantly related animals.

https://royalsocietypublishing.org/doi/pdf/10.1098/rsob.160207

The Drosophila chromosomal protein Mst77F is processed to generate an essential component of mature sperm chromatin.

Inherited microorganisms can selfishly manipulate host reproduction to drive through populations. In Drosophila melanogaster, germline expression of the native Wolbachia prophage WO proteins CifA and CifB cause cytoplasmic incompatibility (CI) in which embryos from infected males and uninfected females suffer catastrophic mitotic defects and lethality; however, in infected females, CifA expression rescues the embryonic lethality and thus imparts a fitness advantage to the maternally-transmitted Wolbachia. Despite widespread relevance to sex determination, evolution, and vector control, the mechanisms underlying when and how CI impairs male reproduction remain unknown and a topic of debate. Here we use cytochemical, microscopic, and transgenic assays in D. melanogaster to demonstrate that CifA and CifB proteins of wMel localize to nuclear DNA throughout the process of spermatogenesis. Cif proteins cause abnormal histone retention in elongating spermatids and protamine deficiency in mature sperms that travel to the female reproductive tract with Cif proteins. Notably, protamine gene knockouts enhance wild type CI. In ovaries, CifA localizes to germ cell nuclei and cytoplasm of early-stage egg chambers, however Cifs are absent in late-stage oocytes and subsequently in fertilized embryos. Finally, CI and rescue are contingent upon a newly annotated CifA bipartite nuclear localization sequence. Together, our results strongly support the Host Modification model of CI in which Cifs initially modify the paternal and maternal gametes to bestow CI-defining embryonic lethality and rescue.

/pdf/the-cif-proteins-from-wolbachia-prophage-wo-modify-sperm-3j8cmofr.pdf

The Cif proteins from Wolbachia prophage WO modify sperm genome integrity to establish cytoplasmic incompatibility

Prtl99C Acts Together with Protamines and Safeguards Male Fertility in Drosophila

Stage-specific testes proteomics of Drosophila melanogaster identifies essential proteins for male fertility.

In the course of spermatogenesis, germ cells undergo dramatic morphological changes that affect almost all cellular components. Therefore, it is impossible to study the process of spermatogenesis in its entirety without detailed morphological analyses. Here, we describe a method to visualize chromatin dynamics in differentiating Drosophila male germ cells using immunofluorescence staining. In addition, we demonstrate how to treat Drosophila sperm before immunofluorescence staining to help reveal epitopes in the highly condensed sperm chromatin that otherwise may be inaccessible to antibodies.

Zeynep Eren-Ghiani

Papers

Prtl99C Acts Together with Protamines and Safeguards Male Fertility in Drosophila

Stage-specific testes proteomics of Drosophila melanogaster identifies essential proteins for male fertility.

Analysis of Chromatin Dynamics During Drosophila Spermatogenesis.