Non-canonical regulation of glycogenolysis and the Warburg phenotype by soluble adenylyl cyclase

Abstract

169/175) 20 21Cyclic AMP is produced in cells by two very different types of adenylyl cyclases: the canonical 22 transmembrane adenylyl cyclases (tmACs, ADCY1~9) and the evolutionarily more conserved soluble 23 adenylyl cyclase (sAC, ADCY10). While the role and regulation of tmACs is well documented, much 24 less is known of sAC in cellular metabolism. We demonstrate here that sAC is an acute regulator of 25 glycolysis, oxidative phosphorylation and glycogen metabolism, tuning their relative bioenergetic 26 contributions. Suppression of sAC activity leads to aerobic glycolysis, enhanced glycogenolysis, 27 decreased oxidative phosphorylation, and an elevated cytosolic NADH/NAD + ratio, resembling the 28 Warburg phenotype. Importantly, we found that glycogen metabolism is regulated in opposite 29 directions by cAMP depending on its location of synthesis and downstream effectors. While the 30 canonical tmAC-cAMP-PKA axis promotes glycogenolysis, we identify a novel sAC-cAMP-Epac1 axis 31 that suppresses glycogenolysis. These data suggest that sAC is an autonomous bioenergetic sensor 32 that suppresses aerobic glycolysis and glycogenolysis when ATP levels suffice. When the ATP level 33 falls, diminished sAC activity induces glycogenolysis and aerobic glycolysis to maintain energy 34 homeostasis.35 36 56 CO2 are the primary product of substrate oxidation by the TCA cycle, we hypothesized that sAC 57 regulates the autonomous metabolism of cells and this is supported by several characteristics of the 58 enzyme. Firstly, sAC can directly sense the cellular energetic state due to its high Km for its substrate 59 ATP (ranging from 1 to 10 mM) (Jaiswal & Conti, 2003, Litvin et al., 2003, Zippin, Chen et al., 2013. 60Secondly, albeit at low level, sAC is expressed in almost all tissues examined (Geng, Wang et al., 2005, 61 Levin & Buck, 2015). Thirdly, Ca 2+ , another versatile and universal second messenger, stimulates sAC 62 in synergy with bicarbonate (Geng et al., 2005, Jaiswal & Conti, 2003, allowing sAC to tune the 63 cellular metabolism according to ongoing cellular signaling. In line with this reasoning, the ATP-and 64 calcium-sensing properties of sAC have been demonstrated to facilitate diverse metabolic 65 regulations, including the regulation of oxidative phosphorylation by sensing CO2 production from 66 the TCA cycle (Acin-Perez, Salazar et al., 2009) and/or free Ca 2+ concentrations in the mitochondrial 67 matrix (Di Benedetto, Scalzotto et al., 2013), induction of ATP-dependent insulin secretion in the 68 pancreas (Zippin et al., 2013), secretion of aldosterone in adrenal cortex carcinoma H295R (Katona, 69 4 Rajki et al., 2015), and TNF-induced respiratory burst in human neutrophils (Han, Stessin et al., 2005). 71Over the past decade, it has become widely accepted that cAMP signaling is compartmentalized 72 into microdomains which allows this single messenger molecule to mediate disparate functions even 73 within a single cell. (Kamenetsky et al., 2006, Lefkimmiatis & Zaccolo, 2014. We have therefore 74 exa...

Cholangiocytes express an isoform of soluble adenylyl cyclase that is N‐linked glycosylated and secreted in extracellular vesicles

“…Glucose deprivation has previously been shown to promote exosome release and modify exosome cargo to proteins related to energy metabolism and related signaling 46 . Given that sAC regulates various cellular glucose‐related responses, 4,47 specific intracellular retention of sAC 75 under glucose deprivation could therefore serve to maintain intrinsic cellular homeostasis.…”

The extracellular lactate-to-pyruvate ratio modulates the sensitivity to oxidative stress-induced apoptosis via the cytosolic NADH/NAD+ redox state

“…Cloning of the Peredox-mCherry NADH/NAD + redox sensor and lentiviral transduction of the sensor into cells was performed as described in [23].…”

“…HepG2 cells stably transduced with the inducible Peredox-mCherry construct or an empty vector (pCW-MCS-BSD) as described in [23]. Cells were induced with 800 ng/mL doxycycline for 48 h prior to the experiment.…”

“…Cells were incubated in HBSS modified for ambient air medium containing a fixed total amount (2.5 mM) of l-lactate plus pyruvate but in different lactate-to-pyruvate ratios (1, 5, 10, 20, 50 and 100) in the presence of glucose. Fluorescence of Peredox (F 1 ) and mCherry (F 2 ) were monitored as described in [23]. Fluorescence from HepG2 cells transduced with the empty vector was used to correct for background fluorescence (denoted as F 1 ′ and F 2 ′).…”