• Effectors Implicated in the Adenylyl Cyclase 1 Inhibitory Effect on Cell Migration in Pancreatic Cancer Cells

      Medepalli, Vidya; Department of Biologic Sciences (2017-03)
      Pancreatic adenocarcinoma is among the most aggressive of all cancers in the United States. Cyclic adenosine monophosphate (cyclic AMP) is a second messenger that regulates the proliferation and migration of pancreatic cancer cells. Cyclic AMP is formed from cytosolic ATP by the enzyme adenylyl cyclase (AC). In the presence of forskolin, a transmembrane AC activator, the proliferation and migration of pancreatic cells have been inhibited. Since the mechanisms underlying the inhibitory effect of activated adenylyl cyclase are little understood, we investigated the downstream mediators implicated in the AC/cyclic AMP pathway. With this purpose, we overexpressed AC1 in the human pancreatic adenocarcinoma (HPAC) cell line, and through the protein kinase A (PKA) inhibitor H-89 and the exchange protein directly activated by cyclic AMP (EPAC) inhibitor ESI-09, we assessed the effector involved upon the treatment with forskolin. In a previous study, we showed that PKA alone mediates the inhibitory effect of forskolin/AC1/cyclic AMP on proliferation of HPAC cells. In the present study, we examined the effectors implicated in the AC1 inhibitory effect on cell migration through utilization of the CytoSelect 24-well cell migration assay kit. Our current experimental data suggests that PKA and EPAC are both likely to be downstream mediators in the effect of forskolin/AC1/cyclic AMP on migration of HPAC cells.
    • The Structural and Functional Properties of a Splice Variant of ADCY7 Gene in Human Pancreatic Cancer

      Liao, Kristie; Jhanji, Nancy; Department of Biological Sciences (2017-03)
      Alternative splicing of mRNA precursors is a regulated process of gene expression in eukaryotic cells. It provides cells with the opportunity to create protein isoforms of different functions from a single gene. Cancer cells often take advantage of this process to produce proteins that promote growth and survival. Cyclic adenosine monophosphate (cyclic AMP) is a second messenger that has shown to suppress migration and invasion of pancreatic ductal adenocarcinoma cells. Cyclic AMP is formed from cytosolic ATP by the enzyme adenylyl cyclase (AC). There are ten isoforms of ACs; nine are anchored in the plasma membrane and one is soluble. Our goal was to find alternative splice variants of transmembrane AC isoforms in pancreatic cancer. We found a possible splice variant of type VII adenylyl cyclase (ADCY7) in human healthy pancreatic, adjacent non-tumor, tumor tissues, two pancreatic cancer cell lines HPAC and PANC-1, epithelial duct cell line PDEC, but not in isolated human pancreatic acini (the exocrine part of the pancreas). Further research will be carried out to study the structural and functional properties of the splice variant of human ADCY7.
    • Structural, Kinetic and Functional Properties of CAP1/AC Complexes

      Mehrotra, Simran; Department of Biological Sciences (2017-03)
      The major cause of death in pancreatic cancer is due to metastases; therefore, it is important to study the mechanism by which the pancreatic cancer cells migrate and invade. This would help advance therapeutics and ultimately help prolong survival. Adenylyl cyclase-associated protein 1 (CAP1) is a scaffold protein that is involved in the regulation of actin microfilament formation, which ultimately leads to cell migration and invasion. CAP1 binds to G-actin inhibiting polymerization. We first tested whether CAP1 binds to adenylyl cyclase (AC) by performing co-immunoprecipitation. We found that CAP1 not only interacts with G-actin, but also with a number of AC isoforms: AC1, AC3, AC4 and AC7. Further studies need to be done to determine how CAP1/AC/G-actin interact and the impact of these interactions on the invasive behavior of pancreatic cancer cells.