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Selective downregulation of human Kv1.3 channels by PKA I
Author(s) -
Kuras Zerrin,
Kucher Vladimir,
Neumeier Lisa,
Dillehay Kelsey,
Conforti Laura
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.24.1_supplement.620.9
Subject(s) - jurkat cells , transfection , downregulation and upregulation , activator (genetics) , hek 293 cells , microbiology and biotechnology , channel blocker , small interfering rna , antagonist , patch clamp , protein kinase a , chemistry , cell culture , agonist , protein subunit , kinase , biology , t cell , receptor , biochemistry , immunology , gene , genetics , immune system , organic chemistry , calcium
The voltage‐gated K channel Kv1.3 plays an important role in the earliest stages of T cell activation and its inhibition suppresses T cell function. Protein kinase A (PKA) modulates Kv1.3 activity and downregulates T cell function. Two types of PKA are expressed in human T cells, Type I and Type II. Molecular and electrophysiological studies were undertaken to determine the PKA isotype modulating Kv1.3. 8Br‐cAMP (8Br), a non‐selective activator of PKA, inhibits Kv1.3 currents by 35% both in primary human T and in Jurkat cells. This inhibition was prevented by the PKA blocker PKI. Jurkat cells were transfected with siRNAs against the regulatory subunit of PKAI and a GFP plasmid (10:1 ratio) for visual recognition during patch‐clamp experiments. Selective downregulation of PKAI knocks down PKAI but not PKAII by 45% as determined by semiquantitative RT‐PCR. 8Br inhibits Kv1.3 current in scramble siRNA transfected (control) cells by 33%. The response to 8Br was abolished in siRNA‐transfected cells. Further experiments in primary T cells showed that the PKAI selective agonist pair 8‐piperidinoadenosine‐cAMP/8‐hexylaminoadenosine‐cAMP induced Kv1.3 inhibition comparable to 8Br, while the PKAI antagonist Rp‐8‐Br‐cAMP significantly reduced 8Br inhibition. These results indicate that PKAI, and not PKAII, is responsible for the regulation of Kv1.3 channels in human T lymphocytes. (NIH AI083076 and AHA 0855457D)