JCS030312 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 -

  Fig.S1. Chaperone recruitment upon expression of mutant vimentin in mice. (A,B) Immunofluorescence of lens specimen stained with antibodies against Hsp70 (red) and total vimentin (green). In lenses of wt animals Hsp70 is hardly detectable, whereas in lenses of VimR₁₁₃C animals Hsp70 is strongly increased and colocalised with vimentin. Bar: 20 µm.

• Supplemental Figure S2 -

  Fig. S2. Unaltered caspase activity and absence of transglutaminase activity in mutant and wt lenses and cells. (A) Luminescent substrate-based proteasome activity assay of extracts from lenses and 3T3L1 fibroblasts stably transfected with VimR₁₁₃C. In both settings, no significant changes in the activity of caspase 3 and caspase 7 were detected. (B) Transglutaminase assay. Lenses of mutant and wt mice were incubated overnight at 37°C in the presence of 2 mM leupeptin. Cell fractions enriched for IF proteins were analysed by SDS-PAGE and immunoblotting. Coomassie Blue staining of the fractions and immunoblot using a vimentin antibody are shown. No crosslinked vimentin was detected in any of the samples. Asterisk marks position at which crosslinked vimentin was expected.