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Functional analysis of the distal region of the third intracellular loop of PROKR2

Xiao-Tao Zhou1, Dan-Na Chen, Zhi-Qun Xie

  • 1Xiangya Hospital, State Key Laboratory of Medical Genetics, Central South University, Changsha, Hunan Province 410078, PR China; Department of Immunology, Xinjiang Medical University, Urumqi, Xinjiang Uygur Autonomous Region 830054, PR China.

Biochemical and Biophysical Research Communications
|August 24, 2013
PubMed

Insights

Mutations in PROKR2 cause hypogonadotropic hypogonadism and Kallmann syndrome. Charged substitutions at residue 274 impair PROKR2 function and cell surface expression, impacting reproductive health.

Area of Science:

  • Endocrinology
  • Molecular Biology
  • Genetics

Background:

  • Mutations in the G-protein-coupled receptor PROKR2 are linked to idiopathic hypogonadotropic hypogonadism (IHH) and Kallmann syndrome (KS), conditions characterized by delayed puberty and infertility.
  • A specific mutation, V274D, was recently found in a patient with KS exhibiting apparent reversal of hypogonadism, prompting further investigation into its mechanism.

Purpose of the Study:

  • To investigate the functional consequences of the V274D mutation and related substitutions (V274A, V274T, V274R) on PROKR2 cell surface expression and signaling activity.
  • To examine the impact of deletions in basic amino acid clusters near residue 274 on PROKR2 trafficking and G-protein coupling.

Main Methods:

  • Site-directed mutagenesis to create V274D, V274A, V274T, V274R, RRK (270-272) deletion, and RKR (264-266) deletion PROKR2 variants.
  • Cell-based assays to assess PROKR2 cell surface expression (e.g., Western blot, immunofluorescence) and signaling activity (e.g., G-protein activation assays).

Main Results:

  • Charged amino acid substitutions at PROKR2 residue 274 (V274D) led to significantly reduced cell surface expression and loss-of-function.
  • Deletion of the RRK motif (270-272) resulted in undetectable PROKR2 cell surface expression.
  • Deletion of the RKR motif (264-266) allowed normal cell surface expression but impaired G-protein coupling, indicating a loss-of-function.

Conclusions:

  • The region around residue 274 in PROKR2, particularly charged substitutions, is critical for proper receptor trafficking and function.
  • Specific basic amino acid clusters within the third intracellular loop (IL3) differentially regulate PROKR2 cell surface expression and G-protein coupling, offering insights into the pathogenesis of IHH and KS.
Keywords:
ERG-protein-coupled receptorGPCRHEKIHHILIntracellular loopKSKallmann syndromeMAPKPK2PROKR2Prokineticin 2Prokineticin receptor 2TMWTendoplasmic reticulumhuman embryonic kidneyidiopathic hypogonadotropic hypogonadismintracellular loopmitogen-activated protein kinasetransmembrane domainwild-type

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