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. 2001 Oct 29;155(3):439–446. doi: 10.1083/jcb.200107114

Figure 1.

Figure 1.

Expression of full-length P0 and P0 deletion mutants in transfected L cells. (A) Diagram of P0 cDNA showing the relative positions of the primers used to clone full-length and mutant P0. The same 5′ primer (P0WT5) was used in all cases. The 3′ primers were designed to allow the cDNAs to be inserted in frame with the flag sequence in the pCMV-Tag4 vector. SP, signal sequence; EC, extracellular domain; TM, transmembrane domain; IC, intracellular domain. (B) P0 mRNA is expressed in L cells transfected with the P0 constructs as determined by RT-PCR using the specific primers shown in A. Primers specific to the enzyme HPRT were used to compare expression levels between cell lines. Numbers to the right of the figure represent the migration of molecular markers. (C) Full-length and mutant P0s are expressed at the cell surface in transfected L cells. Cell membrane proteins were labeled with biotin, and P0 was precipitated with an anti-P0 antibody. The immunoprecipitates were fractionated by SDS-PAGE, transferred to PVDF membranes, and probed with HRP-streptavidin. WT, full-length P0; Δ13, Δ28, Δ43, and Δ59, P0 constructs lacking the terminal 13, 28, 43, or 59 amino acids, respectively; Vect, L cells transfected with empty vector. Numbers to the left of the figure represent the migration of molecular markers.