More than protamines: Identification of further sperm proteins from which Prtl99C is essential for male fertility and full chromatin compaction in Drosophila melanogaster

Spermatogenesis is a highly conserved process that can be divided into three stages: pre-meiosis, meiosis and post-meiosis. At the end of the post-meiotic stage of spermatogenesis, a protamine rich spermatozoon is formed from a histone rich spermatid with round nucleus. During this stage, the chroma...

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Bibliographic Details
Main Author: Eren Ghiani, Zeynep
Contributors: Renkawitz-Pohl, Renate (Prof. Dr.) (Thesis advisor)
Format: Dissertation
Language:English
Published: Philipps-Universität Marburg 2016
Biologie
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Summary:Spermatogenesis is a highly conserved process that can be divided into three stages: pre-meiosis, meiosis and post-meiosis. At the end of the post-meiotic stage of spermatogenesis, a protamine rich spermatozoon is formed from a histone rich spermatid with round nucleus. During this stage, the chromatin is highly packed and the volume of the nucleus is remarkably reduced. Therefore, it is expectable that the poor packed chromatin within the sperm nuclei cannot protect the DNA properly against chemical and physical damage, potentially leading to mutations and infertility problems. In Drosophila, during the replacement process of histones by protamines several high mobility group-box (HMG-box) containing proteins are expressed. Our group previously identified Drosophila protamines being not essential for fertility in contrast to those of mice which cause sterility already in haplo-insufficiency. In both organisms, the compaction of the genome is very efficient despite of the mutated protamine genes. On the basis of these findings, we hypothesized that other proteins may act in functional redundancy or in an additive manner with protamines. For this reason, we searched for additional HMG-box-containing proteins which might play a role during chromatin condensation. We identified Prtl99C specifically expressed in male germ cells. We demonstrated with transgenic flies and with an antibody against Prtl99C that the protein persists until the mature sperm. Analysis of homozygous Prtl99C-ΔC sperm showed longer nuclei, indicating that the C-terminal region is required for chromatin compaction moreover affects male fertility. Knockdown of Prtl99C via RNA interference (RNAi) led also to elongation of the sperm nucleus region by about 35% yet the fertility was highly reduced. Whereas loss of protamines led to an 8% longer nucleus, deletion of both protamines and Prtl99C, led to an elongation of the sperm nucleus by 47%. Based on these findings, we proposed that many chromatin-binding proteins have a function in chromatin reorganization during spermatogenesis in favor of building a compacted male genome. The actual work presents for the first time a Drosophila sperm chromatin component that is essential for male fertility. The insights obtained from this study can help to understand in particular the condensation of the sperm genome carried out by several basic proteins. Lastly, we analyzed a comparative proteome approach to find more basic protein and identified a homolog to mammalian THEG as likely being essential for male fertility Drosophila.
DOI:https://doi.org/10.17192/z2016.0082