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RNAseq for perigonadal WAT of adipose tissue specific CDK7 knockout mice under CL or saline treatment for 7 days.

ABSTRACT: Purpose: The goal of this study are to compare global gene expression profile in pgWAT in Cdk7 aKO and WT control mice after CL or saline treatment for 7 days. Methods: pgWAT pads were taken from 9-week-old CDK7 aKO and control littermates either treated with CL or saline control for 7 days. Mice were sacrificed at zeitgeber time 9. Total RNA was extracted with TRI Reagent (Sigma). Two RNA samples from two mice with same genotype and condition were pool together (each 250 ng) to constitute 500 ng of total RNA from biological triplicates for sequencing. Results: Reads were also aligned to the Mus musculus.GRCm38.86 transcriptome using STAR (v. 2.5.2b, Dobin et al. 2013) and the estimation of the isoforms abundance was computed using RSEM (v. 1.2.31, Li and Dewey 2011). Conclusions: Differentially expressed genes (DEGs) were identified between genotype and treatment.

ORGANISM(S): Mus musculus

PROVIDER: GSE149123 | GEO | 2020/05/20

REPOSITORIES: GEO

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Project description:Luteolysis of the corpus luteum (CL) during non-fertile cycles involves a cessation of progesterone (P4) synthesis (functional regression) and subsequent structural remodeling. The molecular processes responsible for initiation of luteal regression in the primate CL are poorly defined. Therefore, a genomic approach was utilized to systematically identify differentially expressed genes in the rhesus macaque CL during spontaneous luteolysis. CL were collected prior to (days 10-11 post-LH surge, mid-late [ML] stage) or during (days 14-16, late stage) functional regression. Based on P4 levels, late stage CL were subdivided into functional late (FL, serum P4 > 1.5 ng/ml) and functionally-regressed late (FRL, serum P4 < 0.5 ng/ml) groups (n=4 CL/group). Total RNA was isolated, labeled and hybridized to Affymetrix genome microarrays that contain elements representing the entire rhesus macaque transcriptome. With the ML stage serving as the baseline, there were 681 differentially expressed transcripts (>2-fold change; p< 0.05) that could be categorized into three primary patterns of expression: 1) increasing from ML through FRL, 2) decreasing from ML through FRL, and 3) increasing ML to FL, followed by a decrease in FRL. Ontology analysis revealed potential mechanisms and pathways associated with functional and/or structural regression of the macaque CL. Quantitative real-time PCR was used to validate microarray expression patterns of 13 genes with the results being consistent between the two methodologies. Protein levels were found to parallel mRNA profiles in 4 of 5 differentially expressed genes analyzed by Western blot. Thus, this database will facilitate the identification of mechanisms involved in primate luteal regression. Keywords: time course plus functional state of corpus luteum The first experimental factor is the stage of the luteal phase when the CL was collected. There were two stages: 1) between days 10-12 post LH-surge (mid-late stage), and 2) days 14-16 (late stage). The mid-late stage is a transitionary period where CL are still functional (i.e., producing significant quantities of P4) but are nearing the time when luteolysis initiates in non-conception cycles, and the late stage corresponds to the normal period when functional regression (cessation of P4 secretion) occurs in non-conception cycles. The second experimental factor is the functional state of the CL which was determined by measuring serum concentrations of P4 at the time of CL collection. Late stage CL with serum P4 levels > 1.5 ng/ml were considered to be functional, and those with serum P4 < 0.5 ng/ml were considered to have completed functional regression. Measuring mRNA levels in CL collected from these groups provides a comprehensive analysis of the primate transcriptome throughout the normal period of functional regression in the macaque CL. There were 12 biological replicates: n = 4 per group, 3 groups in total. The mid-late CL were used as the baseline reference for gene expression.

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RNAseq for perigonadal WAT of adipose tissue specific CDK7 knockout mice under CL or saline treatment for 7 days.

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