Radiation-Induced RhoGDIβ Cleavage Leads to Perturbation of Cell Polarity: A Possible Link to Cancer Spreading

Abstract

The equilibrium between proliferation and apoptosis is tightly balanced to maintain tissue homeostasis in normal tissues and even in tumors. Achieving and maintaining such a balance is important for cancer regrowth and spreading after cytotoxic treatments. Caspase-3 activation and tumor cell death following anticancer therapy as well as accompanying cell death pathways are well characterized, but their association to homeostasis of cancerous tissue and tumor progression remains poorly understood. Here we proposed a novel mechanism of cancer spreading induced by caspase-3. RhoGDIβ, known as a direct cleavage substrate of caspase-3, is overexpressed in many epithelial cancers. The N-terminal-truncated RhoGDIβ (ΔN-RhoGDIβ) is accumulated in caspase-3-activated cells. Stable expression of ΔN-RhoGDIβ in HeLa cells did not induce apoptosis, but impaired directional cell migration in a wound-healing assay accompanied by a perturbed direction of cell division at the wound edge. Subcellular protein fractionation experiments revealed that ΔN-RhoGDIβ but not wild-type RhoGDIβ was present in the detergent-soluble cytoplasmic and nuclear fractions and preferentially associated with Cdc42. Furthermore, Cdc42 activity was constitutively inhibited by stable expression of ΔN-RhoGDIβ, resulting in increased radiation-induced compensatory proliferation linking to RhoA activation. Thus, ΔN-RhoGDIβ dominant-negatively regulates Cdc42 activity and contributes to loss of polarity-related functions. The caspase-3-cleaved RhoGDIβ is a possible determinant to promote cancer spreading due to deregulation of directional organization of tumor cell population and inhibition of default equilibrium between proliferation and apoptosis after cytotoxic damage. J. Cell. Physiol. 231: 2493-2505, 2016. © 2016 Wiley Periodicals, Inc.

2‐O‐Octadecylascorbic acid represses RhoGDIβ expression and ameliorates DNA damage‐induced abnormal spindle orientations

“…Unfortunately, as the pharmaceutical usefulness might be compromised by the fact that AA itself had no effect on DNA protection and repair in HeLa cells exposed to oxidative stress, 31 we focused on the cellular processes related to compensatory cell division in DNA‐damaged cells. RhoGDIβ is upregulated in apoptosis‐induced compensatory proliferation in HeLa cells 19 ; therefore, we first examined whether AA and its derivatives influenced RhoGDIβ expression.…”

“…HeLa cells irradiated with 10 Gy of X‐rays showed RhoGDIβ upregulation and continuous ΔN‐RhoGDIβ expression for up to 6 days 19 . To determine how long ΔN‐RhoGDIβ expression persisted, the X‐ray‐irradiated cells were cultured for up to 3 weeks.…”

“…Its dysregulation is implicated in the impairment of organ structures and functions, and is related to carcinogenesis 10,30,32–35 . Our previous studies demonstrated that a RhoGDIβ C‐terminal fragment (ΔN‐RhoGDIβ) is persistently expressed in response to X‐irradiation and thereby inhibits Cdc42 activity, which leads to spindle misorientation in the irradiated cells 19 . Further evaluation revealed that shRNA‐mediated knockdown of RhoGDIβ rescued the spindle misorientation caused by radiation exposure 20 .…”

“…This cleavage produces a C‐terminal fragment (N‐terminal‐deleted, ΔN‐RhoGDIβ) that redistributes from the plasma membrane to the cytoplasm and the nucleus 17,18 . We have previously shown perturbed spindle orientation to be associated with upregulated expression of RhoGDIβ and ΔN‐RhoGDIβ in nonpolarized adherent HeLa cells surviving radiation exposure, leading to increased apoptosis‐induced compensatory proliferation 19,20 . These findings highlighted the importance of RhoGDIβ expression on spindle orientation in surviving cells following DNA damage.…”

RhoGDIβ affects HeLa cell spindle orientation following UVC irradiation

“…*p < 0.005 versus HeLa; **p < 10 −5 versus HeLa; ***p < 0.0005 versus HeLa; ****p < 10 −4 versus HeLa. N. S.: not significant; RhoGDIβ: Rho GDP-dissociation inhibitor β; UVC: ultraviolet C [Color figure can be viewed at wileyonlinelibrary.com] cell growth again (Fujiwara et al, 2016). In particular, we focused on RhoGDIβ, which inhibits Rho GTPases in a rheostat-like manner (Boulter & Garcia-Mata, 2010) and is a possible factor that affects spindle orientation, among other caspase-3 substrates (Fujiwara et al, 2016).…”