Spatial alterations of De Novo purine biosynthetic enzymes by Akt-independent PDK1 signaling pathways

Abstract

A macromolecular complex of the enzymes involved in human de novo purine biosynthesis, the purinosome, has been shown to consist of a core assembly to regulate the metabolic activity of the pathway. However, it remains elusive whether the core assembly itself can be selectively controlled in the cytoplasm without promoting the purinosome. Here, we reveal that pharmacological inhibition of the cytoplasmic activity of 3-phosphoinositide-dependent protein kinase 1 (PDK1) selectively promotes the formation of the core assembly, but not the purinosome, in cancer cells. However, alternative signaling cascades that are associated with the plasma membrane-bound PDK1 activity, including Akt-mediated cascades, regulate neither the core assembly nor the purinosome in our conditions. Along with immunofluorescence microscopy and a knock-down study against PDK1 using small interfering RNAs, we reveal that cytoplasmic PDK1-associated signaling pathways regulate subcellular colocalization of three enzymes that form the core assembly of the purinosome in an Akt-independent manner. Collectively, this study reveals a new mode of compartmentalization of purine biosynthetic enzymes controlled by spatially resolved signaling pathways.

Light-sensitive phosphorylation regulates enzyme activity and filament assembly of human IMPDH1 retinal splice variants

“…Purinosome assembly correlates with increased flux through the pathway (Pareek et al, 2020; Zhao et al, 2015) and has been hypothesized to be regulated by posttranslational modifications (Liu et al, 2019). In fact, purinosome assembly is modulated by casein kinase II, AMP-activated protein kinase, and 3-phosphoinositide-dependent protein kinase 1, suggesting various signaling cascades spatially regulate purinosome enzymes (An et al, 2010; Schmitt et al, 2016; Schmitt et al, 2018). It is enticing to speculate that a network of phosphorylation and/or other modifications may spatiotemporally coordinate assembly of IMPDH into various subcellular compartments depending on the metabolic state of the cell.…”

From purines to purinergic signalling: molecular functions and human diseases

“… 32 Moreover, 3-phosphoinositide-dependent protein kinase 1 (PDK1) has been found to regulate the core assembly of purinosome through its cytoplasmic activity, but independent of its membrane-bound activity. 33 It is worth mentioning that G-protein coupled receptor (GPCR) signalling, which regulates a great number of kinases such as PI3K, PKA, Akt, PKC, facilitates purinosome assembly, suggesting that additional kinases are probably involved. 34 …”

Surveying purine biosynthesis across the domains of life unveils promising drug targets in pathogens

“…69 Found in the cytoplasm in association with mitochondria and microtubules, formation is regulated by G-protein coupled receptor signaling, phosphorylation and interaction with molecular chaperones such as HSP90. [70][71][72][73] Purinosomes form primarily, but not exclusively, during the G1 phase of the cell cycle, preceding S phase when purine demand is high because of DNA replication. 74 Unlike gene fusion, compartmentalization of purine biosynthesis is a reversible biomolecular assembly of enzymes that allows short-term regulation in response to metabolic cues.…”

“…The enzymes from the second half of the pathway appear to transiently interact with this core to form the purinosome 69 . Found in the cytoplasm in association with mitochondria and microtubules, formation is regulated by G‐protein coupled receptor signaling, phosphorylation and interaction with molecular chaperones such as HSP90 70‐73 . Purinosomes form primarily, but not exclusively, during the G1 phase of the cell cycle, preceding S phase when purine demand is high because of DNA replication 74 …”