Cdc42‐Borg4‐Septin7 axis regulates HSC polarity and function

Abstract

Aging of hematopoietic stem cells (HSCs) is caused by the elevated activity of the small RhoGTPase Cdc42 and an apolar distribution of proteins. Mechanisms by which Cdc42 activity controls polarity of HSCs are not known. Binder of RhoGTPases proteins (Borgs) are known effector proteins of Cdc42 that are able to regulate the cytoskeletal Septin network. Here, we show that Cdc42 interacts with Borg4, which in turn interacts with Septin7 to regulate the polar distribution of Cdc42, Borg4, and Septin7 within HSCs. Genetic deletion of either Borg4 or Septin7 results in a reduced frequency of HSCs polar for Cdc42 or Borg4 or Septin7, a reduced engraftment potential and decreased lymphoid‐primed multipotent progenitor (LMPP) frequency in the bone marrow. Taken together, our data identify a Cdc42‐Borg4‐Septin7 axis essential for the maintenance of polarity within HSCs and for HSC function and provide a rationale for further investigating the role of Borgs and Septins in the regulation of compartmentalization within stem cells.

Septins in Stem Cells

“…Normal levels of Cdc42 activity (more GDP) regulate the assembly of Septin fibers via "free to bind Borgs", and Borgs lead to a stabilization and bundling of Septin filaments, which is consistent with the strong focal assembly (big blob) of Septins seen in young HSCs (Figure 1B). Such a view is further consistent with the recent finding from our laboratory that upon aging of HSCs, there is a switch in the Cdc42-Borg-Septin relationship: Elevated Cdc42 activity in aged correlated with co-localization of Cdc42 and Borg4, whereas in young HSCs (lower Cdc42 activity) Borg4 and Sept7 co-localized (Kandi et al, 2021) (Figure 1A). In conclusion, these data support that Cdc42 activity influences Septin behavior in both yeast and adult stem cells like HSCs and might also be critical for the role of Septins in quiescence.…”

“…Interestingly though, Septins can be organized in multiple types of structures, implying a versatile but also likely a cell type-specific organization of Septins to serve a special purpose within a cell. Septins show spot-like and blob-like distributions within cells under certain conditions (isolated BD3 domain expression of Borg3 ( Joberty et al, 2001 )) and in certain cell types as well ( Kandi et al, 2021 ) which are likely not linked to fibers. The ring structure of Septins is associated with cytokinesis in yeast, but Septin rings are also formed around internalized bacteria upon infection of mammalian cell lines with Listeria and Shigella bacteria ( Mostowy et al, 2009 ) and intra-cytosolic Shigella are compartmentalized in Septin cage-like structures for autophagy as a host defense ( Mostowy et al, 2010 ).…”

“…In young HSCs (from mice that are 4 months old) there is a polar distribution of the polarity proteins RhoGTPase Cell division control 42 protein (Cdc42), Histone 4 Lysine 16 acetylation (H4K16ac), Per2, Numb, Tubulin and also Sept7, with usually one big blob at one spot with perhaps several smaller spots, while in aged HSCs (from animals 18 months and older), this distribution is apolar: more smaller spots, and a much more dispersed distribution ( Figure 1B ). It is thought that the level of polarity is linked to the mode (symmetric/asymmetric) of stem cell division to balance self-renewal versus differentiation ( Florian et al, 2018 ) and to regulate the homing capability to allow for successful immune system reconstitution after transplantation ( Liang et al, 2005 ; Janzen et al, 2006 ; Florian and Geiger, 2010 ; Dykstra et al, 2011 ; Florian et al, 2012 ; Sun et al, 2014 ; Kandi et al, 2021 ).…”

“…A functional mammalian equivalent was found in 1999 by Joberty et al: Borgs (Binder of Rho GTPases) 1-5, also entitled as Cdc42 effector proteins 1-5, bind Cdc42 in a GTP-dependent manner ( Joberty et al, 1999 ). HSCs do also express Borg proteins ( Kandi et al, 2021 ). Among the above mentioned pathways linked to quiescence, also Iqg1 of the yeast TORC1 pathway was shown to be involved in Septin ring organization and regulation of Cdc42 activity ( Iwase et al, 2007 ; Atkins et al, 2013 ).…”

“…As Septins are master regulators of asymmetry and polarity ( Spiliotis and McMurray, 2020 ), it is very likely that they play a role not only in the asymmetric division of budding yeast but as well also in HSC division and general asymmetric distribution of proteins within HSCs. We could recently demonstrate that Sept7 is indeed involved in regulating the distribution of polarity proteins in HSCs ( Kandi et al, 2021 , see Table1 ). Budding yeast achieves polarity and thus asymmetry during cell cytokinesis by three consecutive steps with enduring Septin support: 1) deciding upon a bud site and 2) passage of cellular components through a barrier and 3) active retainment of components within a compartmentalized space.…”