Cytoplasmic, nuclear, and golgi localization of RGS proteins: Evidence for N-terminal and RGS domain sequences as intracellular targeting motifs

Tapan Kumar Chatterjee, Rory A. Fisher

Research output: Contribution to journalArticle

109 Citations (Scopus)

Abstract

RGS proteins comprise a family of proteins named for their ability to negatively regulate heterotrimeric G protein signaling. Biochemical studies suggest that members of this protein family act as GTPase-activating proteins for certain Gα subunits, thereby accelerating the turn-off mechanism of Gα and terminating signaling by both Gα and Gβγ subunits. In the present study, we used confocal microscopy to examine the intracellular distribution of several RGS proteins in COS-7 cells expressing RGS-green fluorescent protein (GFP) fusion proteins and in cells expressing RGS proteins endogenously. RGS2 and RGS10 accumulated in the nucleus of COS-7 cells transfected with GFP constructs of these proteins. In contrast, RGS4 and RGS16 accumulated in the cytoplasm of COS-7 transfectants. As observed in COS-7 cells, RGS4 exhibited cytoplasmic localization in mouse neuroblastoma cells, and RGS10 exhibited nuclear localization in human glioma cells. Deletion or alanine substitution of an N-terminal leucine repeat motif present in both RGS4 and RGS16, a domain identified as a nuclear export sequence in HIV Rev and other proteins, promoted nuclear localization of these proteins in COS-7 cells. In agreement with this observation, treatment of mouse neuroblastoma cells with leptomycin B to inhibit nuclear protein export by exportin1 resulted in accumulation of RGS4 in the nucleus of these cells. GFP fusions of RGS domains of RGS proteins localized in the nucleus, suggesting that nuclear localization of RGS proteins results from nuclear targeting via RGS domain sequences. RGSZ, which shares with RGS-GAIP a cysteine-rich string in its N-terminal region, localized to the Golgi complex in COS-7 cells. Deletion of the N-terminal domain of RGSZ that includes the cysteine motif promoted nuclear localization of RGSZ. None of the RGS proteins examined were localized at the plasma membrane. These results demonstrate that RGS proteins localize in the nucleus, the cytoplasm, or shuttle between the nucleus and cytoplasm as nucleo-cytoplasmic shuttle proteins. RGS proteins localize differentially within cells as a result of structural differences among these proteins that do not appear to be important determinants for their G protein-regulating activities. These findings suggest involvement of RGS proteins in more complex cellular functions than currently envisioned.

Original languageEnglish (US)
Pages (from-to)24013-24021
Number of pages9
JournalJournal of Biological Chemistry
Volume275
Issue number31
DOIs
StatePublished - Aug 4 2000

Fingerprint

RGS Proteins
COS Cells
Green Fluorescent Proteins
Proteins
Cytoplasm
Cell Nucleus Active Transport
Nuclear Proteins
Neuroblastoma
Cysteine
Fusion reactions
rev Gene Products
Cells
Human Immunodeficiency Virus Proteins
GTPase-Activating Proteins
Heterotrimeric GTP-Binding Proteins
Aptitude
Terminal Repeat Sequences
Confocal microscopy
Golgi Apparatus
Cell membranes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Cytoplasmic, nuclear, and golgi localization of RGS proteins : Evidence for N-terminal and RGS domain sequences as intracellular targeting motifs. / Chatterjee, Tapan Kumar; Fisher, Rory A.

In: Journal of Biological Chemistry, Vol. 275, No. 31, 04.08.2000, p. 24013-24021.

Research output: Contribution to journalArticle

@article{90ae706a931f415eacf0e36f854431ba,
title = "Cytoplasmic, nuclear, and golgi localization of RGS proteins: Evidence for N-terminal and RGS domain sequences as intracellular targeting motifs",
abstract = "RGS proteins comprise a family of proteins named for their ability to negatively regulate heterotrimeric G protein signaling. Biochemical studies suggest that members of this protein family act as GTPase-activating proteins for certain Gα subunits, thereby accelerating the turn-off mechanism of Gα and terminating signaling by both Gα and Gβγ subunits. In the present study, we used confocal microscopy to examine the intracellular distribution of several RGS proteins in COS-7 cells expressing RGS-green fluorescent protein (GFP) fusion proteins and in cells expressing RGS proteins endogenously. RGS2 and RGS10 accumulated in the nucleus of COS-7 cells transfected with GFP constructs of these proteins. In contrast, RGS4 and RGS16 accumulated in the cytoplasm of COS-7 transfectants. As observed in COS-7 cells, RGS4 exhibited cytoplasmic localization in mouse neuroblastoma cells, and RGS10 exhibited nuclear localization in human glioma cells. Deletion or alanine substitution of an N-terminal leucine repeat motif present in both RGS4 and RGS16, a domain identified as a nuclear export sequence in HIV Rev and other proteins, promoted nuclear localization of these proteins in COS-7 cells. In agreement with this observation, treatment of mouse neuroblastoma cells with leptomycin B to inhibit nuclear protein export by exportin1 resulted in accumulation of RGS4 in the nucleus of these cells. GFP fusions of RGS domains of RGS proteins localized in the nucleus, suggesting that nuclear localization of RGS proteins results from nuclear targeting via RGS domain sequences. RGSZ, which shares with RGS-GAIP a cysteine-rich string in its N-terminal region, localized to the Golgi complex in COS-7 cells. Deletion of the N-terminal domain of RGSZ that includes the cysteine motif promoted nuclear localization of RGSZ. None of the RGS proteins examined were localized at the plasma membrane. These results demonstrate that RGS proteins localize in the nucleus, the cytoplasm, or shuttle between the nucleus and cytoplasm as nucleo-cytoplasmic shuttle proteins. RGS proteins localize differentially within cells as a result of structural differences among these proteins that do not appear to be important determinants for their G protein-regulating activities. These findings suggest involvement of RGS proteins in more complex cellular functions than currently envisioned.",
author = "Chatterjee, {Tapan Kumar} and Fisher, {Rory A.}",
year = "2000",
month = "8",
day = "4",
doi = "10.1074/jbc.M002082200",
language = "English (US)",
volume = "275",
pages = "24013--24021",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "31",

}

TY - JOUR

T1 - Cytoplasmic, nuclear, and golgi localization of RGS proteins

T2 - Evidence for N-terminal and RGS domain sequences as intracellular targeting motifs

AU - Chatterjee, Tapan Kumar

AU - Fisher, Rory A.

PY - 2000/8/4

Y1 - 2000/8/4

N2 - RGS proteins comprise a family of proteins named for their ability to negatively regulate heterotrimeric G protein signaling. Biochemical studies suggest that members of this protein family act as GTPase-activating proteins for certain Gα subunits, thereby accelerating the turn-off mechanism of Gα and terminating signaling by both Gα and Gβγ subunits. In the present study, we used confocal microscopy to examine the intracellular distribution of several RGS proteins in COS-7 cells expressing RGS-green fluorescent protein (GFP) fusion proteins and in cells expressing RGS proteins endogenously. RGS2 and RGS10 accumulated in the nucleus of COS-7 cells transfected with GFP constructs of these proteins. In contrast, RGS4 and RGS16 accumulated in the cytoplasm of COS-7 transfectants. As observed in COS-7 cells, RGS4 exhibited cytoplasmic localization in mouse neuroblastoma cells, and RGS10 exhibited nuclear localization in human glioma cells. Deletion or alanine substitution of an N-terminal leucine repeat motif present in both RGS4 and RGS16, a domain identified as a nuclear export sequence in HIV Rev and other proteins, promoted nuclear localization of these proteins in COS-7 cells. In agreement with this observation, treatment of mouse neuroblastoma cells with leptomycin B to inhibit nuclear protein export by exportin1 resulted in accumulation of RGS4 in the nucleus of these cells. GFP fusions of RGS domains of RGS proteins localized in the nucleus, suggesting that nuclear localization of RGS proteins results from nuclear targeting via RGS domain sequences. RGSZ, which shares with RGS-GAIP a cysteine-rich string in its N-terminal region, localized to the Golgi complex in COS-7 cells. Deletion of the N-terminal domain of RGSZ that includes the cysteine motif promoted nuclear localization of RGSZ. None of the RGS proteins examined were localized at the plasma membrane. These results demonstrate that RGS proteins localize in the nucleus, the cytoplasm, or shuttle between the nucleus and cytoplasm as nucleo-cytoplasmic shuttle proteins. RGS proteins localize differentially within cells as a result of structural differences among these proteins that do not appear to be important determinants for their G protein-regulating activities. These findings suggest involvement of RGS proteins in more complex cellular functions than currently envisioned.

AB - RGS proteins comprise a family of proteins named for their ability to negatively regulate heterotrimeric G protein signaling. Biochemical studies suggest that members of this protein family act as GTPase-activating proteins for certain Gα subunits, thereby accelerating the turn-off mechanism of Gα and terminating signaling by both Gα and Gβγ subunits. In the present study, we used confocal microscopy to examine the intracellular distribution of several RGS proteins in COS-7 cells expressing RGS-green fluorescent protein (GFP) fusion proteins and in cells expressing RGS proteins endogenously. RGS2 and RGS10 accumulated in the nucleus of COS-7 cells transfected with GFP constructs of these proteins. In contrast, RGS4 and RGS16 accumulated in the cytoplasm of COS-7 transfectants. As observed in COS-7 cells, RGS4 exhibited cytoplasmic localization in mouse neuroblastoma cells, and RGS10 exhibited nuclear localization in human glioma cells. Deletion or alanine substitution of an N-terminal leucine repeat motif present in both RGS4 and RGS16, a domain identified as a nuclear export sequence in HIV Rev and other proteins, promoted nuclear localization of these proteins in COS-7 cells. In agreement with this observation, treatment of mouse neuroblastoma cells with leptomycin B to inhibit nuclear protein export by exportin1 resulted in accumulation of RGS4 in the nucleus of these cells. GFP fusions of RGS domains of RGS proteins localized in the nucleus, suggesting that nuclear localization of RGS proteins results from nuclear targeting via RGS domain sequences. RGSZ, which shares with RGS-GAIP a cysteine-rich string in its N-terminal region, localized to the Golgi complex in COS-7 cells. Deletion of the N-terminal domain of RGSZ that includes the cysteine motif promoted nuclear localization of RGSZ. None of the RGS proteins examined were localized at the plasma membrane. These results demonstrate that RGS proteins localize in the nucleus, the cytoplasm, or shuttle between the nucleus and cytoplasm as nucleo-cytoplasmic shuttle proteins. RGS proteins localize differentially within cells as a result of structural differences among these proteins that do not appear to be important determinants for their G protein-regulating activities. These findings suggest involvement of RGS proteins in more complex cellular functions than currently envisioned.

UR - http://www.scopus.com/inward/record.url?scp=0034604693&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034604693&partnerID=8YFLogxK

U2 - 10.1074/jbc.M002082200

DO - 10.1074/jbc.M002082200

M3 - Article

C2 - 10791963

AN - SCOPUS:0034604693

VL - 275

SP - 24013

EP - 24021

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 31

ER -