Unknown,Transcriptomics,Genomics,Proteomics

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Loss of the RNA-binding protein Kep1 leads to cytokinesis defects and behavioural abnormalities in male Drosophila.

ABSTRACT: The kep1 gene encodes an RNA-binding protein containing a single maxi-KH domain. We have previously demonstrated that loss of the kep1 gene results in Drosophila females displaying a reduction in fertility and wished to further characterize the kep1 mutation in Drosophila males. Wild-type females mated to homozygous kep1- males resulted in 6% of the eggs laid hatching. This reduced reproductive success is due in part to a meiosis defect during spermatogenesis with approximately 10 % of sperm demonstrating a defect in cytokinesis. Utilizing indirect immunohistochemistry we found that the Kep1 protein is present in the nucleus of adult brain neuronal cells, suggesting a behavioural component contributing to the almost complete male sterility phenotype. We report that homozygote kep1- males display almost no courtship behaviour with a measured median courtship index of 0.005 relative to a median index of 0.77 for OreR wild-type controls. In order to better understand the behavioural role of kep1, we carried out gene profiling experiments to identify transcripts whose steady-state levels are altered in the kep1- homozygote males. Our gene profiling studies identified 61 transcripts whose steady-state levels are altered by at least 2 fold or more comparing RNA isolated from w1118 and kep1-/kep1- male heads. A large percentage of transcripts identified whose steady-state expression levels are depleted in kep1-/kep1- heads (10 out of 44) encode for proteins involved in some aspect of proteolysis. Our results show that the Kep1 protein has a pleiotropic effect in Drosophila males leading to cytokinesis defects and behavioural abnormalities. RNA was isolated from heads of 3-4 day old wild-type or homozygote mutant males. Three indepentent RNA samples were utilized to complete experiment.

ORGANISM(S): Drosophila melanogaster

SUBMITTER: Marco Di Fruscio

PROVIDER: E-GEOD-6086 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Although the evidence for a genetic predisposition to human essential hypertension is compelling, the genetic control of blood pressure (BP) is poorly understood. The Dahl salt-sensitive (S) rat is a model for studying the genetic component of BP. Using this model we previously reported the identification of 16 different genomic regions that contain one or more BP quantitative trait loci (QTLs). The proximal region of rat chromosome 1 contains multiple BP QTLs. Of these, we have localized the BP QTL1b region to a 13.5cM (20Mb) region. Interestingly, five additional independent studies in rats and four independent studies in humans have reported genetic linkage for BP control by regions homologous to QTL1b. To view the overall renal transcriptional topography of the positional candidate genes for this QTL, we sought a comparative gene expression profiling between a congenic strain containing QTL1b and control S rats by employing: (1) a saturated QTL1b interval specific oligonucleotide array, and (2) a whole genome cDNA microarray representing 20,465 unique genes that are positioned outside the QTL. Results indicated that 19 out of the 231 positional candidate genes for this QTL are differentially expressed between the two strains tested. Surprisingly, over 1,500 genes outside of QTL1b were differentially expressed between the two rat strains. Integrating the results from the two approaches revealed at least one complex network of transcriptional control initiated by the positional candidate Nr2f2. This network appears to account for the majority of gene expression differences occurring outside of the QTL interval. Further substitution mapping is currently underway to test the validity of each of these differentially expressed positional candidate genes. These results demonstrate the importance of using a saturated oligonucleotide array for identifying and prioritizing differentially expressed positional candidate genes of a BP QTL. Pairs of Cy5 and Cy3 labeled targets were co-hybridized onto either the oligonucleotide microarray or a custom TIGR rat cDNA array consisting of 26,401 probe elements representing 20,465 unique non-QTL1b genes. A âflip-dyeâ design was used as the experimental method of choice to account for potential dye-bias labeling effects. Seven âflip dyeâ normalized files are submitted for the oligonucleotide array and twelve individual hybridizations are submitted for the cDNA array.

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Loss of the RNA-binding protein Kep1 leads to cytokinesis defects and behavioural abnormalities in male Drosophila.

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