DNA-PKcs-mediated phosphorylation of AMPKγ1 regulates lysosomal AMPK activation by LKB1

Abstract

Autophagy is a central component of the cytoprotective cellular stress response. To enlighten stress-induced autophagy signaling, we screened a human kinome siRNA library for regulators of autophagic flux in MCF7 human breast carcinoma cells and identified the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) as a positive regulator of basal and DNA damage-induced autophagy. Analysis of autophagy-regulating signaling cascades placed DNA-PKcs upstream of the AMP-dependent protein kinase (AMPK) and ULK1 kinase. In normal culture conditions, DNA-PKcs interacted with AMPK and phosphorylated its nucleotide-sensing g1 subunit at Ser-192 and Thr-284, both events being significantly reduced upon AMPK activation. Alanine substitutions of DNA-PKcs phosphorylation sites in AMPKg1 reduced AMPK activation without affecting its nucleotide sensing capacity. Instead, the disturbance of DNA-PKcs-mediated phosphorylation of AMPKg1 inhibited the lysosomal localization of the AMPK complex and its starvationinduced association with LKB1. Taken together, our data suggest that DNA-PKcs-mediated phosphorylation of AMPKg1 primes AMPK complex to the lysosomal activation by LKB1 thereby linking DNA damage response to autophagy and cellular metabolism.

DNA-PKcs interacts with and phosphorylates Fis1 to induce mitochondrial fragmentation in tubular cells during acute kidney injury

“…In addition to the nucleus, it can also localize to the cytoplasm where it controls cytoskeleton rearrangement ( 14 ), apoptosis initiation ( 9 ), protein ubiquitination ( 39 ), and autophagy activation ( 40 ), among other cellular processes. DNA-PKcs can localize to multiple organelles, including mitochondria ( 17 ), the Golgi apparatus ( 13 ), and lysosomes ( 41 ). In the Golgi, doxorubicin treatment or oxidative stress mediates the translocation of DNA-PKcs from the nucleus into the cytoplasm where DNA-PKcs interacts with and induces Golgi dispersal and cell death by promoting GOLPH3 phosphorylation ( 13 ).…”

“…In the Golgi, doxorubicin treatment or oxidative stress mediates the translocation of DNA-PKcs from the nucleus into the cytoplasm where DNA-PKcs interacts with and induces Golgi dispersal and cell death by promoting GOLPH3 phosphorylation ( 13 ). During autophagy activation, DNA-PKcs phosphorylates the nucleotide-sensing γ1 subunit of AMPK1 ( 41 ), which correlates with a decrease in starvation-induced autophagy. In aged mice, DNA-PKcs activation represses AMPK phosphorylation ( 17 ), thereby suppressing mitochondrial biogenesis and a decline in the number of mitochondria.…”

Loss of Prom1 impairs autophagy and promotes epithelial‐mesenchymal transition in mouse retinal pigment epithelial cells

“…In addition, Prom1-KO inhibits NUAK1 gene expression in mRPE cells (Figure 4e). NUAK1, an AMPK-related kinase (also known as Ark5), increases the cellular bioenergetic status through mitochondrial respiration (Escalona et al, 2020) and is a regulator of autophagic flux (Puustinen et al, 2020). These observations demonstrate that Prom1 plays a critical role in maintaining mRPE lysosomal function, proteolytic activity, and bioenergetics through TFEB activation.…”

DNA-PKcs: A Targetable Protumorigenic Protein Kinase

“…Nevertheless, DNA-PKcs inhibition or knockdown has been shown to promotes apoptosis and sensitize cells in response to heat shock (134), ultrasound (135), and anti-cancer agents such as etoposide and doxorubicin (6,136). DNA-PKcs has also been shown promote autophagy in cancer through regulation of AMPK in response to etoposide (137) and IR (138). Additionally, DNA-PKcs modulates senescence in response to IR in cancer (139).…”