5 research outputs found
The metabolic enzyme hexokinase 2 localizes to the nucleus in AML and normal haematopoietic stem and progenitor cells to maintain stemness
Thomas, Egan et al. report that hexokinase 2 localizes to the nucleus of leukaemic and normal haematopoietic cells to maintain stemness by interacting with nuclear proteins and modulating chromatin accessibility independently of its kinase activity. Mitochondrial metabolites regulate leukaemic and normal stem cells by affecting epigenetic marks. How mitochondrial enzymes localize to the nucleus to control stem cell function is less understood. We discovered that the mitochondrial metabolic enzyme hexokinase 2 (HK2) localizes to the nucleus in leukaemic and normal haematopoietic stem cells. Overexpression of nuclear HK2 increases leukaemic stem cell properties and decreases differentiation, whereas selective nuclear HK2 knockdown promotes differentiation and decreases stem cell function. Nuclear HK2 localization is phosphorylation-dependent, requires active import and export, and regulates differentiation independently of its enzymatic activity. HK2 interacts with nuclear proteins regulating chromatin openness, increasing chromatin accessibilities at leukaemic stem cell-positive signature and DNA-repair sites. Nuclear HK2 overexpression decreases double-strand breaks and confers chemoresistance, which may contribute to the mechanism by which leukaemic stem cells resist DNA-damaging agents. Thus, we describe a non-canonical mechanism by which mitochondrial enzymes influence stem cell function independently of their metabolic function
Human SAS‑6 C‑Terminus Nucleates and Promotes Microtubule Assembly <i>in Vitro</i> by Binding to Microtubules
Centrioles
are essential components of the animal centrosome and play crucial
roles in the formation of cilia and flagella. They are cylindrical
structures composed of nine triplet microtubules organized around
a central cartwheel. Recent studies have identified spindle assembly
abnormal protein SAS-6 as a critical component necessary for formation
of the cartwheel. However, the molecular details of how the cartwheel
participates in centriolar microtubule assembly have not been clearly
understood. In this report, we show that the C-terminal tail (residues
470–657) of human SAS-6, HsSAS-6 C, the region that has been
shown to extend toward the centriolar wall where the microtubule triplets
are organized, nucleated and induced microtubule polymerization <i>in vitro</i>. The N-terminus (residues 1–166) of HsSAS-6,
the domain known to be involved in formation of the central hub of
the cartwheel, did not, however, exert any effect on microtubule polymerization.
HsSAS-6 C bound to the microtubules and localized along the lengths
of the microtubules <i>in vitro</i>. Microtubule pull-down
and coimmunoprecipitation (Co-IP) experiments with S-phase synchronized
HeLa cell lysates showed that the endogenous HsSAS-6 coprecipitated
with the microtubules, and it mediated interaction with tubulin. Isothermal
calorimetry titration and size exclusion chromatography showed that
HsSAS-6 C bound to the αβ-tubulin dimer <i>in vitro</i>. The results demonstrate that HsSAS-6 possesses an intrinsic microtubule
assembly promoting activity and further implicate that its outer exposed
C-terminal tail may play critical roles in microtubule assembly and
stabilizing microtubule attachment with the centriolar cartwheel