A Human Tyrosine Phosphatase Interactome Mapped by Proteomic Profiling.

Tyrosine phosphatases play a critical role in many cellular processes and pathogenesis, yet comprehensive analysis of their functional interacting proteins in the cell is limited. By utilizing a proteomic approach, here we present an interaction network of 81 human tyrosine phosphatases built on 1884 high-confidence interactions of which 85% are unreported. Our analysis has linked several phosphatases with new cellular processes and unveiled protein interactions genetically linked to various human diseases including cancer. We validated the functional importance of an identified interaction network by characterizing a distinct novel interaction between PTPN5 and Mob1a. PTPN5 dephosphorylates Mob1a at Y26 residue. Further, we identify that PTPN5 is required for proper midbody abscission during cytokinesis through regulation of Mob1a dephosphorylation. In conclusion, our study provides a valuable resource of tyrosine phosphatase interactions, which can be further utilized to dissect novel cellular functions of these enzymes.

The site of action of the antiterminator protein N from the lambdoid phage H-19B.

Transcription antitermination by N proteins of lambdoid phages involves specific interactions of the C-terminal domain of N with the elongation complex (EC). The interacting surface of N on the EC is unknown, knowledge of which is essential to understand the mechanism of antitermination. Specific cleavage patterns were generated near the active site Mg2+ of the RNA polymerase of an N-modified stalled EC using iron-(S)-1-(p-bromoacetamidobenzyl)ethylenediaminetetraacetate conjugated to the only cysteine residue in the C-terminal domain of N from a lambdoid phage H-19B. Modification of EC by N also induced conformational changes around the same region as revealed from the limited trypsin digestion and in situ Fe-dithiothreitol cleavage pattern of the same EC. These data, together with the previously obtained H-19B N-specific mutations in RNA polymerase, beta (G1045D), and beta' (P251S, P254L, G336S, and R270C) subunits, suggest that the active center cleft of the EC could be the site of action of this antiterminator. H-19B N induced altered interactions in this region of EC, prevented the backtracking of the stalled EC at the ops pause site and destabilized RNA hairpin-beta subunit flap domain interactions at the his pause site. We propose that the physical proximity of the C-terminal domain of H-19B N to the active center cleft of the EC is required for the process of transcription antitermination and that it involves both stabilization of the weak RNA-DNA hybrid at a terminator and destabilization of the interactions of terminator hairpin in the RNA exit channel.