Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Comparison of testis transcriptome changes resulting from mutations in dany and representative meiotic arrest genes

ABSTRACT: The Drosophila gene dany is expressed specifically in spermatocytes and encodes a nuclear protein with similarity to the products of the genes distal antenna (dan) and distal antenna-related (danr). Loss of dany function results in male sterility. Mutant spermatocytes fail to differentiate similar as previously observed in mutants lacking tTAF and tMAC function. Microarray analyses were performed to characterize the effects of loss of dany function on gene expression in testis and for a comparison of the effects with those caused by mutations in the tTAF gene spermatocyte arrest (sa) and the tMAC genes always early (aly) and matotopetli (topi) RNA was isolated after testis dissection from adult males and used for the production of probes for microarray hybridization. Five different genotypes were used for testis dissection: control, dany mutant, sa mutant, aly mutant, topi mutant. The number of biological replicates per genotypes was between two and four.

ORGANISM(S): Drosophila melanogaster

SUBMITTER: Christian Lehner

PROVIDER: E-GEOD-76298 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Drosophila dany is essential for transcriptional control and nuclear architecture in spermatocytes.

Trost Martina M Blattner Ariane C AC Leo Stefano S Lehner Christian F CF

Development (Cambridge, England) 20160701 14

The terminal differentiation of adult stem cell progeny depends on transcriptional control. A dramatic change in gene expression programs accompanies the transition from proliferating spermatogonia to postmitotic spermatocytes, which prepare for meiosis and subsequent spermiogenesis. More than a thousand spermatocyte-specific genes are transcriptionally activated in early Drosophila spermatocytes. Here we describe the identification and initial characterization of dany, a gene required in sperma ...[more]

PMID: 27436041

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Project description:The piRNA pathway is studied in great detail in Drosophila female germline. In this study we show that unlike the female germline where all Piwi proteins are expressed throughout oogenesis, Ago3 - a Piwi family protein shows a spatial expression male germline. To understand dynamics of piRNA pathway during spermatogonia and primary spermatocyte stages of male germline development, we used arrest mutants. The bag of marbles (bam) and benign gonial cell neoplasm (bgcn) mutants have only early mitotic dividing germline cells in the testes due to failure to progress to primary spermatocyte stage, the cannonball (can) and spermatocyte arrest (sa) mutant germline cells cannot progress beyond primary spermatocyte stage. To investigate the dynamics of the piRNA pathway during spermatogenesis in spermatogonia and primary spermatocyte stages, we used testicular tissues from these stage-specific arrested mutants. While we used entire bam and bgcn mutant testes for spermatogonia purification, we while we manually removed the apical regions of can and sa mutant testes to exclude mitotically dividing undifferentiated germline cells for primary spermatocytes purification. Our results show that piRNAs mapping to transposons are more abundant in spermatogonia, whereas those mapping to Suppressor of Stellate [Su(Ste)] and AT-chX are mostly expressed in primary spermatocytes. Furthermore we observed that transposon-mapping piRNAs with ping-pong signature are more abundant in spermatogonia albeit still detectable in primary spermatocytes where Ago3 is not expressed. These results suggest that robust piRNA production via ping-pong cycle takes place in spermatogonia, and to a lesser extent in primary spermatocytes even in the absence of Ago3. Consistently, piRNAs from ago3 mutant testes also exhibit the ping-pong signature, confirming that a non-canonical ping-pong cycle is acting during spermatogenesis. Our study provides a developmental dimension to the piRNA pathway and uncovers a new mechanism used in the male germline to silence transposons. The difference in piRNA from spermatogonia and primary spermatocyte stages was studied by comparing small RNAs from bam and bgcn mutant testis, which represent spermatogonia stages with the small RNAs from apex removed can and sa testis, representing primary spermatocyte stages. In the study we also studied effect of loss of Piwi family proteins Aub and Ago3, which have different spatial expression during male germline development.

Project description:The role of different proteins, Always Early (Aly), Spermatocyte Arrest (Sa), Ubi-p63E (Magn) on the gene expression in spermatocyte differentation was assessed by microarray ABSTRACT: The ubiquitin proteasome system (UPS) regulates many biological pathways by posttranslationally ubiquitinating proteins for degradation. Although maintaining a dynamic balance between free ubiquitin and ubiquitinated proteins is key to UPS function, the mechanisms that regulate ubiquitin homeostasis in different tissues through development are not clear. Here we show that loss of function of Drosophila magellan (magn), the polyubiquitin Ubi-p63E, results in specifically meiotic arrest sterility in males. Expression of ubiquitin from magn/Ubi-p63E contributes predominantly to maintaining the free ubiquitin pool in testes. Function of magn/Ubip63E is required cell autonomously for proper meiotic chromatin condensation, cell cycle progression and spermatid differentiation. magn/Ubi-p63E mutant germ cells develop normally to the spermatocyte stage but arrest at the G2/M transition of meiosis I with lack of protein expression of key meiotic cell cycle regulators Boule and Cyclin B. Loss of function of magn/Ubi-p63E did not strongly affect the spermatocyte transcription program regulated by the tTAF and tMAC genes. Knocking down proteasome function specifically in spermatocytes caused a different meiotic arrest phenotype, suggesting that the magn/Ubi-p63E phenotype may not result from general defects in protein degradation. Our results suggest a conserved role of polyubiquitin genes in male meiosis and a potential mechanism leading to meiosis I maturation arrest. RNA was obtained from whole testes of flies with following mutations: 1) aly[2]/aly[5p], 2) sa[1]/sa[2], 3) magn[12c]/magn[23b] or magn[12c]/magn[12c];pSC2, 4) red[1], e[1] (WT). Each genotype had three biological replicates except magn which had four total biological replicates, two from magn[12c]/[23b] and two from [12c]/[12c]; pSC2

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Dataset Information

Comparison of testis transcriptome changes resulting from mutations in dany and representative meiotic arrest genes

Publications

Drosophila dany is essential for transcriptional control and nuclear architecture in spermatocytes.

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