Congratulations Nick Martinez (Campbell lab) on his first authored paper in Journal of Molecular Biology!
Summary
The KRAS/PI3Ka听interaction is a crucial interaction in cancer, but the only available structural information is of the RAS/PI3Kg听complex. Isolated binding domains of RAS effectors are commonly used for biophysical and structural studies of RAS/effector interactions due to difficulties in generating functional full-length effector proteins. PI3K is difficult to generate, produced in low yield and with variable activity, so we generated and characterized functional RAS binding domains (RBDs) for PI3Ka听and PI3K驳听that associate with GTP-RAS with similar affinity to full-length PI3K isoforms. These novel PI3K RBDs will facilitate characterization of RAS/PI3K interactions and provide new insights into how PI3K isoforms differentially recognize RAS. This knowledge could help expedite drug discovery efforts that aim to inhibit the KRAS/PI3Ka听颈苍迟别谤补肠迟颈辞苍.
Abstract
Phosphatidylinositol-3-kinases (PI3Ks) are lipid kinases that phosphorylate听phosphatidylinositol 4,5-bisphosphate to generate a key lipid second messenger, phosphatidylinositol 3,4,5-bisphosphate.听 PI3K补听and听PI3K驳听require activation by RAS proteins to stimulate signaling pathways that control听cellular growth, differentiation, motility and survival.听Intriguingly, RAS binding to PI3K isoforms likely differ, as RAS mutations have been identified that discriminate between PI3K补听and听PI3Kg,听consistent with low sequence homology (23%) between their RAS binding domains (RBDs). As disruption of the RAS/PI3Ka听interaction reduces tumor growth in mice with RAS- and epidermal growth factor receptor driven skin and lung cancers, compounds that interfere with this key interaction may prove useful as anti-cancer agents.听 However, a structure of PI3Ka听bound to RAS is lacking, limiting drug discovery efforts. Expression of full-length PI3K isoforms in insect cells has resulted in low yield and variable activity, limiting biophysical and structural studies of RAS/PI3K interactions.听 This led us to generate the first RBDs from PI3Ka听and PI3Kg听that can be expressed at high yield in bacteria and bind to RAS with similar affinity to full-length PI3K.听 We also solved a 2.31听脜 X-ray crystal structure of the听PI3Ka-RBD, which aligns well to full-length PI3Ka. Structural differences between the PI3Ka听and PI3Kg听RBDs are consistent with differences in thermal stability and may underly differential RAS recognition and RAS-mediated PI3K activation. These high expression, functional PI3K RBDs will aid in interrogating RAS interactions and could aid in identifying inhibitors of this key interaction.
About Nicholas G. Martinez
After initially pursuing a career as a K-3rd grade elementary school teacher with additional training and coursework in special education and ESL, Nick Martinez switched careers paths to pursue his dream of being a cancer researcher. Using his mother’s passing from colon cancer at an early age as motivation, he gained a strong academic and research background in biochemistry and structural biology at the University of Kansas and through an NSF REU summer research fellowship in 2016 at Brandeis University. Nick Martinez chose to continue his graduate education at the University of North Carolina at Chapel Hill, where he is currently pursuing a Ph.D. in Biochemistry & Molecular Biology in Dr. Sharon Campbell’s lab.
He is currently a 4th听year BCBP graduate student with an expected graduation date is December 2021. Nick will be joining Dr. Georgios Skiniotis鈥� lab at Stanford University as a postdoctoral fellow to gain cryo-EM experience prior to heading into industry.
All authors: Nicholas G. Martinez, David F. Thieker, Leiah M. Carey, Juhi A. Rasquinha, Samantha K. Kistler, Brian A. Kuhlman, Sharon L. Campbell. Department of Biochemistry & Biophysics, 大象传媒 at Chapel Hill and Lineberger Comprehensive Cancer Center, 大象传媒 at Chapel Hill.