BEGIN:VCALENDAR VERSION:2.0 PRODID:-//Talks.cam//talks.cam.ac.uk// X-WR-CALNAME:Talks.cam BEGIN:VEVENT SUMMARY:Kendrew lecture- Post-translational regulation of cell signalling - Tony Hunter DTSTART:20160525T140000Z DTEND:20160525T150000Z UID:TALK62741@talks.cam.ac.uk CONTACT:Scientific Meetings Co-ordinator DESCRIPTION:Posttranslational modification (PTM) is a means of increasing the complexity of the proteome\, and reversible PTMs are commonly used in the transmission of signals within cells in response to external stimuli. Protein phosphorylation is involved in the majority of cellular processes and thousands of distinct phosphorylation events can be detected in a sing le cell type. The human kinome comprises >530 protein kinases of which 480 are typical eukaryotic protein kinases (ePKs)\, and the remainder are aty pical protein kinases (aPKs)\; the majority are Ser/Thr kinases\, but ther e are 90 Tyr kinases. In addition to Ser\, Thr and Tyr\, six other amino a cids can be phosphorylated\, including the three basic amino acids\, His\, Lys and Arg. Histidine phosphorylation of proteins is becoming increasing ly relevant as a regulatory mechanism\, and two aPKs\, NME1 and NME2\, can reportedly phosphorylate histidine in target proteins. To study global hi stidine phosphorylation events we generated monoclonal antibodies to non-c leavable 1-pHis and 3-pHis analogues\, and are using these mAbs to study h istidine phosphorylation in normal and transformed cells and in metastasis . By using immobilized mAbs for affinity purification of pHis proteins fro m 293 cells\, we have identified ~800 proteins that bound selectively unde r denaturing conditions\, and putatively contain 1-pHis (~250) or 3-pHis ( ~160). We have begun to characterize some of these pHis proteins\, and de velop MS methods to pinpoint the exact sites of His phosphorylation. Immun ostaining of HeLa cells and primary macrophages with anti-1-pHis mAbs reve aled staining on the outside of phagocytic vesicles\; immunostaining of pr oliferating HeLa cells with anti-3-pHis mAbs showed a striking pattern in mitotic cells\, with strong staining of spindle poles (and centrosomes in interphase cells) and the midbody\, suggesting that phosphorylation of His at the 3-position may regulate some aspects of the mitotic process.\n \ nAberrant protein phosphorylation plays a role in many human diseases\, an d particularly in cancer. In this connection\, we have studied the phenoty pes of several newly identified cancer-associated protein kinase mutations \, and have found\, surprisingly\, that mutations in DAPK3\, MLK4\, and pr otein kinase C are all loss of function\, implying that they act as tumor suppressors. The tumor microenvironment\, which comprises many different s tromal cells types in addition to tumor cells\, is now recognized as playi ng a key role in the development of cancer. Pancreatic adenocarcinoma (PDA C) is one of the most deadly cancers\, and PDAC has an unusually dense str omal matrix surrounding the tumor cells. To better understand the relation ship between stromal cells and tumor cells in PDAC\, we have investigated the crosstalk between pancreatic cancer cells and pancreatic stromal cells \, known as stellate cells\, by proteomic analysis of secreted proteins an d of tyrosine phosphorylation events induced by secreted proteins in both cell types. We have found that LIF and IL-6 cytokines secreted by stellat e stromal cells elicit JAK/STAT3 signaling in the tumor cells\, and that P DGF secreted by the tumor cells in turn activates the stellate cells\, thu s setting up a reciprocal paracrine loop that could in principle be target ed for PDAC therapy. As a proof of principle that this is possible\, usin g a mouse model of PDAC\, we have found that administration of a neutraliz ing anti-LIF mAb prolongs survival when used in combination with gemcitabi ne. LOCATION:Max Perutz Lecture Theatre\, Medical Research Council (MRC) (MRC Laboratory of Molecular Biol END:VEVENT END:VCALENDAR