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Adriana E.M Klomp, Bart van de Sluis, Leo W.J Klomp, Cisca Wijmenga 

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Presentation on theme: "Adriana E.M Klomp, Bart van de Sluis, Leo W.J Klomp, Cisca Wijmenga "— Presentation transcript:

1 The ubiquitously expressed MURR1 protein is absent in canine copper toxicosis 
Adriana E.M Klomp, Bart van de Sluis, Leo W.J Klomp, Cisca Wijmenga  Journal of Hepatology  Volume 39, Issue 5, Pages (November 2003) DOI: /S (03)

2 Fig. 1 MURR1 is deleted in CT Bedlington terriers. (A) Immunoblot analysis of MBP and MBP/MURR1. Recombinant proteins (100 ng) were separated by SDS–PAGE, and transferred to nitrocellulose. Proteins were visualized with preimmune serum or anti-MURR1, in some cases preincubated with MBP or MBP/MURR1, all followed by goat anti-rabbit peroxidase conjugate. (B) MURR1 expression in a liver homogenate of a wild-type mouse, a CT-affected Bedlington terrier, or a healthy beagle. Proteins were visualized as in (A). Journal of Hepatology  , DOI: ( /S (03) )

3 Fig. 2 MURR1 is ubiquitously expressed. Immunoblot analysis of MURR1 on different murine tissues samples (A), or cell lysates (B). Proteins were separated by SDS–PAGE, and transferred to nitrocellulose. Proteins were visualized with anti-MURR1 (upper) or anti-SCHAD (lower), followed by goat anti-rabbit peroxidase conjugate. Journal of Hepatology  , DOI: ( /S (03) )

4 Fig. 3 MURR1 protein expression is not influenced by copper levels. HeLa cells, H441 cells, A549 cells (A) or Caco2 cells (A,B) were cultured in the presence of the copper chelator BCS or CuCl2. Cells were lysed and MURR1 and SCHAD protein expression were investigated by immunoblot analysis using anti-MURR1 and anti-SCHAD, respectively, as described in Fig. 1A. (C) Caco2 cells were cultured in the presence of BCS or CuCl2. Total RNA was isolated and the expression of MTIIA mRNA levels was determined by northern blot analysis, using a probe encoding the complete MTIIA cDNA. Journal of Hepatology  , DOI: ( /S (03) )

5 Fig. 4 Subcellular localization of MURR1 using cell fractionation. Total cell lysate was divided into a cytosolic and a membranous fraction. Protein samples were separated by SDS–PAGE, and transferred to nitrocellulose. MURR1 was visualized with anti-MURR1 (upper), followed by goat anti-rabbit peroxidase conjugate. The membrane-bound protein E-cadherin (middle) and the cytosolic protein β-actin (lower) were visualized with monoclonal antibodies against these proteins, followed with goat anti-mouse peroxidase conjugate. Journal of Hepatology  , DOI: ( /S (03) )

6 Fig. 5 Subcellular localization of MURR1 using indirect immunofluorescence. (A) HeLa cells were analyzed by indirect confocal immunofluorescence using preimmune serum, anti-MURR1, anti-MURR1 preincubated with MBP/MURR1, or anti-MURR1 preincubated with MBP, followed by Cy3-conjugated donkey anti-rabbit IgG. (B) HeLa cells were analyzed by double-label indirect immunofluorescence of MURR1 and p230 (upper), TFR (middle), or CD63 (lower), visualized by Cy3-conjugated donkey anti-rabbit IgG and FITC-conjugated donkey anti-mouse IgG, respectively. Overlap in staining patterns is depicted in yellow in the merged images. The depicted scale bars represent 10 μM. Journal of Hepatology  , DOI: ( /S (03) )


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