Leukocyte analysis from WHIM syndrome patients reveals a pivotal role for GRK3 in CXCR4 signaling
Karl Balabanian, … , Fernando Arenzana-Seisdedos, Françoise Bachelerie
Karl Balabanian, … , Fernando Arenzana-Seisdedos, Françoise Bachelerie
Published February 14, 2008
Citation Information: J Clin Invest. 2008;118(3):1074-1084. https://doi.org/10.1172/JCI33187.
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Research Article Immunology

Leukocyte analysis from WHIM syndrome patients reveals a pivotal role for GRK3 in CXCR4 signaling

Abstract

Leukocytes from individuals with warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome, a rare immunodeficiency, and bearing a wild-type CXCR4 ORF (WHIMWT) display impaired CXCR4 internalization and desensitization upon exposure to CXCL12. The resulting enhanced CXCR4-dependent responses, including chemotaxis, probably impair leukocyte trafficking and account for the immunohematologic clinical manifestations of WHIM syndrome. We provided here evidence that GPCR kinase-3 (GRK3) specifically regulates CXCL12-promoted internalization and desensitization of CXCR4. GRK3-silenced control cells displayed altered CXCR4 attenuation and enhanced chemotaxis, as did WHIMWT cells. These findings identified GRK3 as a negative regulator of CXCL12-induced chemotaxis and as a candidate responsible for CXCR4 dysfunction in WHIMWT leukocytes. Consistent with this, we showed that GRK3 overexpression in both leukocytes and skin fibroblasts from 2 unrelated WHIMWT patients restored CXCL12-induced internalization and desensitization of CXCR4 and normalized chemotaxis. Moreover, we found in cells derived from one patient a profound and selective decrease in GRK3 products that probably resulted from defective mRNA synthesis. Taken together, these results have revealed a pivotal role for GRK3 in regulating CXCR4 attenuation and have provided a mechanistic link between the GRK3 pathway and the CXCR4-related WHIMWT disorder.

Authors

Karl Balabanian, Angélique Levoye, Lysiane Klemm, Bernard Lagane, Olivier Hermine, Julie Harriague, Françoise Baleux, Fernando Arenzana-Seisdedos, Françoise Bachelerie

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Figure 7

Expression and stability of GRK3 mRNAs in WHIMWT fibroblasts.

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Expression and stability of GRK3 mRNAs in WHIMWT fibroblasts. ...
Fibroblasts from P3, P4, and healthy subjects were incubated with 10 μg/ml ActD (treatment) for the indicated times (A and B) or for 4 h and further cultured for 4 h in the absence of ActD (chase) (C). At the indicated times, total RNA was extracted and semi-quantitative PCRs were performed using specific primers flanking the full LDH or GRK3 ORF. Amplified cDNA products were run on 1% agarose gels, detected by ethidium bromide staining, and quantified by computed-assisted densitometry using the ImageJ 1.34 software (NIH). Results are from 1 representative experiment out of 3 (A and B) or are means ± SD of 3 independent determinations (C) and indicate the amount of mRNAs that remained after incubation with ActD (A and B) or that accumulated in the course of ActD chase (C). Transcript levels in fibroblasts incubated in medium alone were set as 100%. Kinetics of LDH mRNA appearance were comparable in all cell types (data not shown). *P < 0.05.