Autophagy Reports (Dec 2023)
Transcriptional pausing factor M1BP regulates cellular homeostasis by suppressing autophagy and apoptosis in Drosophila eye
Abstract
During organogenesis cellular homeostasis plays a crucial role in patterning and growth. The role of promoter proximal pausing of RNA polymerase II, which regulates transcription of several developmental genes by GAGA factor or Motif 1 Binding Protein (M1BP), has not been fully understood in cellular homeostasis. Earlier, we reported that M1BP, a functional homolog of ZKSCAN3, regulates wingless (wg) and caspase-dependent cell death (apoptosis) in the Drosophila eye. Further, blocking apoptosis does not fully rescue the M1BPRNAi phenotype of reduced eye. Therefore, we looked for other possible mechanism(s). In a forward genetic screen, members of the Jun-amino-terminal-(NH2)-Kinase (JNK) pathway were identified. Downregulation of M1BP ectopically induces JNK, a pro-death pathway, known to activate both apoptosis and caspase-independent (autophagy) cell death. Activation of JNK pathway components can enhance M1BPRNAi phenotype and vice-versa. Downregulation of M1BP ectopically induced JNK signaling, which leads to apoptosis and autophagy. Apoptosis and autophagy are regulated independently by their genetic circuitry. Here, we found that blocking either apoptosis or autophagy alone rescues the reduced eye phenotype of M1BP downregulation; whereas, blocking both apoptosis and autophagy together significantly rescues the M1BP reduced eye phenotype to near wild-type in nearly 85% progeny. This data suggests that the cellular homeostasis response demonstrated by two independent cell death mechanisms, apoptosis and autophagy, can be regulated by a common transcriptional pausing mechanism orchestrated by M1BP. Since these fundamental processes are conserved in higher organisms, this novel functional link between M1BP and regulation of both apoptosis and autophagy can be extrapolated to humans. Abbreviations: hid: head involution defective; rpr: reaper; DIAP: Drosophila inhibitor of apoptosis proteins; RD genes: Retinal Determination genes; MF: Morphogenetic Furrow; PCD: Programmed cell death; ER: Endoplasmic reticulum; Wg: Wingless; JNK - c: Jun amino-terminal (NH2) Kinase; MAPKs – Mitogen: activated protein kinases; TNF: Tumor necrosis factor; Egr: Eiger; Wgn: Wengen; Tak 1: TGFβ activating kinase 1; JNKKK: JNK kinase kinase; Hep: Hemipterous; JNKK: JNK kinase; Bsk: Basket; Puc - Puckered; Pol II - RNA polymerase II; DNA: Deoxyribonucleic acid; TSS: Transcription start site; ESCs: Embryonic stem cells; Hop: Hopscotch; Dpp: Decapentaplegic; GAF: GAGA factor; M1BP: Motif 1 Binding Protein; ZAD: Zinc-associated domain; ZKSCAN3 - Zinc finger with a SCAN and a KRAB domain 3 (ZKSCAN3); UAS: Upstream activation sequence; ELAV: Embryonic lethal abnormal vision; RT qPCR: Real time quantitative polymerase chain reaction; LOF: Loss of function; GOF: Gain of function; GFP: Green fluorescent protein; WT: Wild-type; PBS: Phosphate buffered saline; PFA: Paraformaldehyde; PBST: Phosphate Buffered Saline with Triton X-100; β-GAL: Beta-galactosidase; DSHB: Developmental Studies Hybridoma Bank; CST: Cell Signaling Technologies; Dlg: Discs large; IgG: Immunoglobulin G; FITC: Fluorescein isothiocyanate; RNA: Ribonucleic acid; cDNA: Complementary DNA; Ct: cycle threshold; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; SAPK: Stress-activated protein kinases; HRP: Horse Radish Peroxidase; ROI: Region Of Interest; SEM: Standard Error of the Mean; CI: Confidence Intervals
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