Downregulation of two novel genes in Sl/Sld and W(LacZ)/Wv mouse jejunum.
Interstitial cells of Cajal (ICC) are the so-called pacemaker cells of the gut. W(LacZ)/Wv and Sl/Sld mice lack ICC surrounding the myenteric plexus (MP) in the jejunum. We compared the gene expression profile of wild type (WT) and W(LacZ)/Wv and Sl/Sld mice using suppression subtractive hybridization (SSH), generating a cDNA library of 1303 clones from which 48 unique sequences were differentially expressed with Southern blot. Among them, we identified heme oxygenase2, TROY, and phospholamban in ICC using immunohistochemistry. Using RT-qPCR, c-Kit and two new transcripts Dithp and prenylcysteine oxidase1 were significantly lower expressed in Sl/Sld and W(LacZ)/Wv versus WT. Prenylcysteine oxidase1 appeared cytotoxic for COS-7 cells and was highly expressed in liver while Dithp was mainly expressed in small intestine. The combination of SSH, Southern blot, RT-qPCR, and immunohistochemistry turned out to be a useful approach to identify rarely expressed genes and genes with small differences in expression.
Wouters
Mira M
MM
Department of Internal Medicine, Johnson and Johnson, Pharmaceutical Research and Development, A Subdivision of Janssen Pharmaceutics, Beerse, Belgium.
Neefs
Jean-Marc
JM
Kerchove d'Exaerde
Alban de
Ad
Vanderwinden
Jean-Marie
JM
Smans
Karine A
KA
D000818:Animals; D019556:COS Cells; D002135:Calcium-Binding Proteins / Q000096:biosynthesis; D013437:Carbon-Sulfur Lyases / Q000096:biosynthesis*; D002522:Chlorocebus aethiops; D015536:Down-Regulation; D020869:Gene Expression Profiling; D006419:Heme Oxygenase (Decyclizing) / Q000096:biosynthesis; D007150:Immunohistochemistry; D007421:Intestine, Small / Q000378:metabolism; D007583:Jejunum / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D019009:Proto-Oncogene Proteins c-kit / Q000096:biosynthesis; D018124:Receptors, Tumor Necrosis Factor / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000099211:Phospholamban
D002135:Calcium-Binding Proteins; D013437:Carbon-Sulfur Lyases; D006419:Heme Oxygenase (Decyclizing); D019009:Proto-Oncogene Proteins c-kit; D018124:Receptors, Tumor Necrosis Factor; D000099211:Phospholamban; C409471:Tnfrsf19 protein, mouse; C105245:heme oxygenase-2; C491543:Pcyox1 protein, mouse