Cloning and characterization of a new intestinal inflammation-associated colonic epithelial Ste20-related protein kinase isoform.
Intestinal epithelial cells respond to inflammatory extracellular stimuli by activating mitogen activated protein kinase (MAPK) signaling, which mediates numerous pathophysiological effects, including intestinal inflammation. Here, we show that a novel isoform of SPS1-related proline alanine-rich kinase (SPAK/STE20) is involved in this inflammatory signaling cascade. We cloned and characterized a SPAK isoform from inflamed colon tissue, and found that this SPAK isoform lacked the characteristic PAPA box and alphaF loop found in SPAK. Based on genomic sequence analysis the lack of PAPA box and alphaF loop in colonic SPAK isoform was the result of specific splicing that affect exon 1 and exon 7 of the SPAK gene. The SPAK isoform was found in inflamed and non-inflamed colon tissues as well as Caco2-BBE cells, but not in other tissues, such as liver, spleen, brain, prostate and kidney. In vitro analyses demonstrated that the SPAK isoform possessed serine/threonine kinase activity, which could be abolished by a substitution of isoleucine for the lysine at position 34 in the ATP-binding site of the catalytic domain. Treatment of Caco2-BBE cells with the pro-inflammatory cytokine, interferon gamma, induced expression of the SPAK isoform. Over-expression of the SPAK isoform in Caco2-BBE cells led to nuclear translocation of an N-terminal fragment of the SPAK isoform, as well as activation of p38 MAP kinase signaling cascades and increased intestinal barrier permeability. These findings collectively suggest that pro-inflammatory cytokine signaling may induce expression of this novel SPAK isoform in intestinal epithelia, triggering the signaling cascades that govern intestinal inflammation.
Yan
Y
Y
Department of Medicine, Division of Digestive Diseases, Emory University School of Medicine, 615 Michael Street, Atlanta, GA 30322, USA.
Nguyen
H
H
Dalmasso
G
G
Sitaraman
S V
SV
Merlin
D
D
D000595:Amino Acid Sequence; D001483:Base Sequence; D018938:Caco-2 Cells; D003001:Cloning, Molecular; D003092:Colitis / Q000201:enzymology* / Q000235:genetics; D003106:Colon / Q000201:enzymology; D006801:Humans; D007413:Intestinal Mucosa / Q000201:enzymology*; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism; D000099317:Hippo Kinases
D009419:Nerve Tissue Proteins; D020033:Protein Isoforms; D017346:Protein Serine-Threonine Kinases; D048051:p38 Mitogen-Activated Protein Kinases; C109163:STK24 protein, human; D000099317:Hippo Kinases; C486409:STK39 protein, human