yHSC AThi oHSC ATlo oHSC Bulk RNA-seq - Understanding transcriptional changes between autophagy engaging and non-engaging hematopoietic stem cells during mouse aging.

We sought to understand the transcriptional profile of autophagy engaging vs non-engaging old hematopoietic stem cells (HSC) referenced against young HSCs. Bulk RNA-seq analyses indicated a large transcriptional divergence between the two oHSC subsets, with pathway analyses of differentially expressed genes (DEG) demonstrating enrichment in inflammatory signaling in AThi oHSCs and in oxidative metabolism signaling in ATlo oHSCs. Ingenuity Pathway Analysis (IPA) inferred that several inflammation-coupled pro-autophagy regulators including p53, Bnip3l, Nupr1, and Nlrp3[22,23], in addition to quiescence enforcing checkpoints FoxO3a, Rb1 and Cdkn1a, were activated in AThi oHSCs. Overall design: RNA from HSCs of two young (8-12wks) and three old GFP-LC3 (>24mo) mice was processed for bulk RNA-sequencing. For the old HSC biological replicates, the 33% GFPlo (AThi) and 33% GFPhi (ATlo) HSCs were isolated by FACS prior to RNA isolation.