Western analysis of PTPROt proteins, human homologues of murine PTPØ. PTPROt cDNAs, which included or lacked bp 680 to 763 from the juxtamembrane region (PTPROt_long[L] and PTPROt_short[S], Fig 2) were in vitro translated, immunoblotted and analyzed with an antiserum directed against the putative murine homologue of PTPROt, PTPØ.8Cell lysates from DHL-4 cells transfected with vector only, pcDNA3-PTPROt_[S]sense, or pcDNA-PTPROt_[S]antisense were similarly immunoblotted and analyzed. As indicated, the predicted ∼47-kD and 43-kD PTPROt_[L] and PTPROt_[S] proteins were readily detectable in in vitro translations of PTPROt sense cDNAs and absent in in vitro translations of PTPROt antisense cDNAs. In the in vitro translated products, the less intense bands running ≈4 kD lower than the major proteins are likely to result from the use of a second start codon (nucleotide 604) with a strong Kozak consensus sequence. DHL-4 PTPROt_[S]sense transfectants also expressed high levels of the expected ≈43-kD protein, which was not detected in DHL-4 PTPROt_[S]antisense transfectants. (The PTPØ antisera also identified ≈70-kD proteins of uncertain significance in all [vector-only, antisense, sense] DHL-4 clones.) The filters were also probed with an antitubulin antibody to assure equal loading.