RGS14 Is a Centrosomal and Nuclear Cytoplasmic Shuttling Protein That Traffics to Promyelocytic Leukemia Nuclear Bodies Following Heat Shock

Abstract

RGS14, a member of the regulator of G-protein signaling (RGS) protein family, possesses an N-terminal RGS domain, two Raf-like Ras-binding domains, and a GoLoco motif, which has GDP dissociation inhibitor activity. In this study we show that unique among the known mammalian RGS proteins, RGS14 localizes in centrosomes. Its first Ras-binding domain is sufficient to target RGS14 to centrosomes. RGS14 also shuttles between the cytoplasm and nucleus, and its nuclear export depends on the CRM-1 nuclear export receptor. Mutation of a nuclear export signal or treatment with leptomycin B causes nuclear accumulation of RGS14 and its association with promyelocytic leukemia protein nuclear bodies. Furthermore, a point mutant defective in nuclear export fails to target to centrosomes, suggesting that nuclear cytoplasmic shuttling is necessary for its proper localization. Mild heat stress, but not proteotoxic or transcription-linked stresses, re-localizes the RGS14 from the cytoplasm to promyelocytic leukemia nuclear bodies. Expression of RGS14, but not point mutants that disrupt the functional activity of its RGS domain or GoLoco motif, enhances the reporter gene activity. The multifunctional domains and the dynamic subcellular localization of RGS14 implicate it in a diverse set of cellular processes including centrosome and nuclear functions and stress-induced signaling pathways. Many of the members of the regulator of G protein signaling (RGS)1 family of over 30 proteins regulate signaling pathways initiated by receptors that couple to heterotrimeric G proteins. They do so by enhancing the rate of GTP hydrolysis of G␣ subunits, thus reducing the duration that the G␣ subunits are GTP-bound (1-4), by antagonizing the interactions between activated G␣ subunits and effectors (2), by directly inhibiting the activities of certain effectors (5, 6), and by inhibiting the dissociation of guanine nucleotides from the G␣ subunits (7). However, emerging evidence suggests that RGS proteins, particularly those that contain domains in addition to their signature RGS domain, have other cellular functions (for recent reviews see Refs. 8 and 9).RGS14 is a GTPase-activating protein (GAP) for the G i ␣ subfamily but not for the G q ␣, G s ␣, or G 12/13 ␣ subfamily (10). It possesses an N-terminal RGS domain that is necessary and sufficient for its GAP activity and several other domains, including two Raf-like Ras-binding domains (RBD) and a GoLoco motif (7, 11). A region overlapping the first RBD domain and portion of the second RBD has been designated as a Rapinteraction domain (RID) because it mediated the interaction of RGS14 with GTP-bound forms of Rap1 and Rap2 but not Ras (12). The functional importance of this binding and whether the RBD domains might interact with other small GTPases is unknown. The GoLoco motif of RGS14 has GDP dissociation inhibitor activity (GDI) for G i ␣. Although its physiological significance remains enigmatic, the GoLoco motif does enhance the inhibitory effect of RGS14 on G i ␣ signaling triggered through th...

Non-visual Arrestins Are Constitutively Associated with the Centrosome and Regulate Centrosome Function

“…Moreover, fluorescence resonance energy transfer experiments reveal that G␣ i1 directly interacts with RGS14 in the centrosome, and its normal expression and function is essential for proper cytokinesis (66). Indeed, silencing G␣ i or RGS14 expression produced cells with multiple nuclei (55), mirroring the defects observed after loss of arrestin2 or -3 expression.…”

“…For example, the sphingosine 1-phosphate receptor (65), G␣ i (66), G␤␥, and RGS14 (55) have been shown to co-localize with ␥-tubulin and/or pericentrin. Moreover, fluorescence resonance energy transfer experiments reveal that G␣ i1 directly interacts with RGS14 in the centrosome, and its normal expression and function is essential for proper cytokinesis (66).…”

Differential expression of regulator of G‐protein signaling R12 subfamily members during mouse development

“…Here, we examined the subcellular distribution of RGS12 splice variants in vivo. Whereas RGS14 is localized to the nucleus and cytoplasm in numerous cell types (Martin-McCaffrey et al, 2004;Cho et al, 2005), RGS12 was localized in the cytoplasm (Fig. 5).…”

Canonical and Noncanonical G-Protein Signaling Helps Coordinate Actin Dynamics To Promote Macrophage Phagocytosis of Zymosan

“…For CXCR4-YFP, YFP was fused to the C-terminus of human CXCR. Elmo1-GFP (kindly provided by Kodi S. Ravichandran), Ags3-RFP (22), AGS4-GFP and LGN-GFP (kindly provided by Joe Blumer), ½ YFP-G␤1 and ½ YFP-G␥2 (kindly provided by Catherine Berlot), Rgs14-GFP (23), and Ric-8A-GFP and Ric-8A-mCherry (24) have been described elsewhere. RAW 264.7 cells were also permanently transfected (via electroporation) with Elmo1-GFP-and EEtagged G␣ i2 QL (Missouri S&T cDNA Resource Center).…”