PT  - JOURNAL ARTICLE
AU  - Maria Guaita-Cespedes
AU  - Rubén Grillo-Risco
AU  - Marta R. Hidalgo
AU  - Sonia Fernández-Veledo
AU  - Deborah Jane Burks
AU  - María de la Iglesia-Vayá
AU  - Amparo Galán
AU  - Francisco Garcia-Garcia
TI  - Assessment of Sex Bias in Housekeeping Gene Expression in Adipose Tissue Through the Massive Analysis of Transcriptomics Data
AID  - 10.1101/2021.12.04.471124
DP  - 2022 Jan 01
TA  - bioRxiv
PG  - 2021.12.04.471124
4099  - http://biorxiv.org/content/early/2022/04/05/2021.12.04.471124.short
4100  - http://biorxiv.org/content/early/2022/04/05/2021.12.04.471124.full
AB  - As the housekeeping genes (HKGs) generally involved in maintaining essential cell functions are typically assumed to exhibit constant expression levels across cell types, they are commonly employed as internal controls in gene expression studies. Nevertheless, multiple studies indicate that not all HKGs display stable expression across cells and tissues and under various healthy and diseased conditions, which can introduce systematic errors into experimental results. The selection and validation of HKGs as controls for each studied condition represent crucial steps in ensuring the validity of obtained results; however, up till now, sex has not been typically considered as a biological variable.In this study, we evaluate the expression profiles of six classical HKGs (four metabolic: GAPDH, HPRT, PPIA, and UBC, and two ribosomal: 18S and RPL19) to determine expression stability in adipose tissues of Homo sapiens and Mus musculus and check sex bias and their overall suitability as internal controls. We also assess the stabile expression of genes included in distinct whole-transcriptome microarrays available from the Gene Expression Omnibus database (GEO) to identify sex-unbiased HKGs suitable for use as internal controls. We perform a sex-based analysis to describe any sexual dimorphisms in mRNA expression stability.A novel computational strategy based on meta-analysis techniques proves that certain classical HKGs fail to function adequately as controls when analyzing human adipose tissue (HAT) considering sex as a variable. The extensively used 18S gene displays sex-based variability in adipose tissue, although PPIA and RPL19 do not, and hence, represent robust HKGs. We propose new sex-unbiased human and mouse HKGs (suHKG) derived from sex-specific expression profiles, such as RPS8 and UBB. All results generated during this study are readily available by accessing an open web resource (https://bioinfo.cipf.es/metafun-HKG) for consultation and reuse in further studies.Competing Interest StatementThe authors have declared no competing interest.