Polo-like kinase 1 directs the AMPK-mediated activation of myosin regulatory light chain at the cytokinetic cleavage furrow independently of energy balance

Abstract

It has been recently proposed that AMP-activated protein kinase (AMPK) might indirectly promote the phosphorylation of MRLC (myosin II regulatory light chain) at Ser19 to regulate the transition from metaphase to anaphase and the completion of cytokinesis. Although these findings provide biochemical support for our earlier observations showing that the active form of the α catalytic AMPK subunit associates dynamically with essential mitotic regulators, several important issues remained unexplored. Does glucose starvation alter the ability of AMPK to bind to the mitotic apparatus and travel from centrosomes to the spindle midzone during mitosis and cytokinesis? Does AMPK activate MRLC exclusively at the cleavage furrow during cytokinesis? What is the mitosis-specific stimulus that activates the mito-cytokinetic AMPK/MRLC axis regardless of energy deprivation? First, we confirm that exogenous glucose deprivation fails to alter the previously described distribution of phospho-AMPKα(Thr172) in all of the mitotic phases and does not disrupt its apparent association with the mitotic spindle and other structures involved in cell division. Second, we establish for the first time that phospho-AMPKα(Thr172) colocalizes exclusively with Ser19-phosphorylated MRLC at the cleavage furrow of dividing cells, a previously unvisualized interaction between phospho-AMPKα(Thr172) and phospho-MRLC(Ser19) that occurs in cleavage furrows, intercellular bridges and the midbody during cell division that appears to occur irrespective of glucose availability. Third, we reveal for the first time that the inhibition of AMPK mitotic activity in response to PLK1 inhibition completely prevents the co-localization of phospho-AMPKα(Thr172) and phospho-MRLC(Ser19) during the final stages of cytokinesis and midbody ring formation. Because PLK1 inhibition efficiently suppresses the AMPK-mediated activation of MRLC at the cytokinetic cleavage furrow, we propose a previously unrecognized role for AMPK in ensuring that cytokinesis occurs at the proper place and time by establishing a molecular dialog between PLK1 and MRLC in an energy-independent manner.

Ser2481-autophosphorylated mTOR colocalizes with chromosomal passenger proteins during mammalian cell cytokinesis

“…Using an automated confocal imaging system for high-resolution images and 3-D reconstructions, we first explored the spatiotemporal dynamics of phospho-mTOR Ser2481 during the G 2 /M stage of the cell cycle in HeLa cancer cells. Similar to that observed when analyzing the spatiotemporal dynamics of phospho-AMPK Thr172 during mitosis and cytokinesis, [18][19][20][21][22][23] we observed a previously unrecognized pattern of transient associations between phosphomTOR Ser2481 and specific mitotic structures during mitotic chromosome condensation/segregation and cytokinesis, including the central spindle midzone and the midbody (Fig. 2).…”

“…All spindle midzone and the midbody, throughout all of the mitotic stages and during the furrowing process in cytokinesis. [18][19][20][21][22][23] Indeed, a trimeric complex of AMPK containing the γ 2 regulatory subunit appears to become selectively activated in the cytokinetic apparatus and to be directly linked to mitotic processes. 22,24,25 Other groups have confirmed these observations, [26][27][28] and we now know that mitotic AMPK regulates mitotic spindle orientation through phosphorylation of the myosin regulatory light chain, and that it ensures the normal recruitment of myosin molecules into the contractile ring structure to allow the proper transition from metaphase to anaphase and the completion of cytokinesis.…”

Role of SVIL phosphorylation by PLK1 in myosin II activation and cytokinetic furrowing

“…Rho-associated kinase (ROCK) phosphorylates RLC to initiate constriction of the contractile ring, and PLK1 can phosphorylate ROCK to promote catalytic activity during cytokinesis (Lowery et al, 2007). A recent study showed that PLK1 phosphorylates the energy-sensing AMPactivated protein kinase (AMPK) to promote phosphorylation of RLC at the furrow (Vazquez-Martin et al, 2012). In addition to these pathways, our study showed that PLK1 can regulate myosin II activation by phosphorylating SVIL.…”

The PTEN-Akt pathway impacts the integrity and composition of mitotic centrosomes

“…During interphase, Akt and AMPK have antagonistic effects on the mTOR pathway and subsequently on cell progression. However, the nutrient-sensing functions of AMPK are dispensable for its mitotic functions, 41 and thus PLK-1 may act as a bridge to connect the AMPK and Akt pathways to ensure successful completion of mitosis. The ability of Akt and PTEN to regulate centrosome composition during mitosis likely contributes to proper spindle formation, as demonstrated by the increase in centrosome defects caused by the knockdown of PTEN and/ or inhibition of Akt with MK2206 (Fig.…”