Identification of SRSF9 through pooled shRNA screening links BNIP3 splicing to autophagy and metabolic reprogramming in breast cancer

Autophagy is a catabolic process that facilitates the degradation of toxic molecules and contributes to the maintenance of cellular homeostasis. However, the alternative splicing of genes involved in hypoxia-induced autophagy remains relatively unexplored. Here we performed pooled shRNA screening of multiple RNA binding proteins under hypoxia to mechanistically decipher their role in regulating alternative splicing of autophagy genes in breast cancer cells. We reported reduced SRSF9 expression under hypoxia that when exogenously expressed diminishes autophagosome formation. SRSF9 regulates alternative splicing of BNIP3 generating two functionally distinct isoforms regulating autophagosome formation and cancer progression through Warburg effect. Mechanistically, SRSF9 directly binds to exon 3 of BNIP3 mRNA inhibiting the truncated BNIP3- Δ3 generation and resultant warburg effect. Moreover, the individual ectopic expression of full length BNIP-FL promotes autophagosome formation. Collectively, our study identifies novel mechanism where SRSF9 mediates the switch between autophagy and Warburg effect via BNIP3 alternative splicing thereby impacting breast cancer progression and chemoresistance.