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Figure 4


Fig. 4. Region II of Nvj1p sequesters Tsc13p. (A) The membrane-spanning domain of Nvj1p (region II) is responsible for recruiting Tsc13p-EYFP. Full-length (FL) or N- and C-terminal truncations of Nvj1p were overexpressed in nvj1-{Delta} cells and tested for the ability to sequester Tsc13p-EYFP from cortical ER compartments to the perinuclear ER. Nuclear chromatin (blue) and vacuole membranes (red) were co-stained with Hoechst and FM4-64, respectively. Regions flanking either side of the membrane-spanning domain of Nvj1p (denoted by a shaded box) proved dispensable for the sequestration of Tsc13p-EYFP to the nuclear envelope. Numbers correspond to amino acid positions. (B) The transmembrane region of Nvj1p (aa 85-120) is sufficient to co-immunoprecipitate Tsc13p-EYFP in vivo. Detergent-extracted lysates of cells expressing Tsc13p-EYFP and 3HA-tagged Nvj1p(85-120aa) or empty vector were prepared as described in Materials and Methods. Nvj1p(85-120aa)-3HA was immunoprecipitated (IP) with anti-HA conjugated agarose beads.