Differences in intracellular signaling patterns in HUVECs initially attaching to various ECM component combinations

Abstract

Program/Abstract # 50Differences in intracellular signaling patterns in HUVECs initiallyattaching to various ECM component combinationsChristine Pauken, Michael CaplanSBHSE, Arizona State University, Tempe, AZ, USAWhen tissues are growing, injured, or in some pathological states,the vasculature must remodel itself in response to the new micro-environments. The new microenvironments include different compo-nents of the extracellular matrix (ECM) that the endothelial cells areexposed to and these cells must decipher and respond to the newcomponents. In this study, HUVECs were placed on 15 differentcombinationsoftwoECMcomponentsfor5–360 min.ThelevelsofErk1/2, Jnk, Akt, and NFKB phosphorylationwere measured at eight timepointsoverthisperiod.The480datapointswereanalyzedbyprincipalcomponent analysis.Thefirst principalcomponent correlateswithErk1/2signalingintheHUVECs,andthescoreplotshowshighcorrelationswithCol1/IV,ColIV/Lm,Vn/CSandVn/HS.FurtherstatisticalanalysisshowsthatdifferentErk1/2signalingpatternsoccurinHUVECsonCol1/Col IV, Col 1/Lm, Col IV/Lm, Col IV/HS, Vn/HS, and Vn/CS. OtherchangesinsignalingpatternsareseenintheJnkandAktphosphoryla-tionpatternswhentheHUVECsareplacedontheseECMcombinations.This data will enhance the knowledge of how HUVECs interpret theirmicroenvironment and allow for a better understanding of woundrepair, vascularization in tumors, and improved design of biomaterialsfor blood vessel repair and tissue engineering.doi:10.1016/j.ydbio.2010.05.083Program/Abstract # 51A role for Mxtx2 in mesendoderm inductionBenjamin Feldman, Sung-Kook Hong, Jamie L. BrownMedical Genetics Branch, National Human Genome Research Institute,NIH, Bethesda, MD 20892, USAA long-standing challenge to embryologists is deciphering theinstructions guiding germ layer differentiation at the onset ofgastrulation. In vertebrates, Nodal signaling is a core component ofmesoderm and endoderm (mesendoderm) differentiation and muchremains to be learned about how expression of the Nodal ligandsthemselves is activated. Motivated byevidence that the zebrafishyolkharbors Nodal- and mesendoderm-activating signals, we developed amicrodissection strategy for the genome-wide identification oftranscripts enriched in the early yolk. This analysis led us to reassessthe function of Mxtx2, a transcription factor that has been shown byothers to be expressed in the yolk and to be essential for normalepiboly. We find that Mxtx2 is a strong inducer of mesendoderm andits loss of function causes substantial reductions in the earlyexpression of the Nodal-related genes and several other mesendo-derm genes. We further find that Mxtx2 directly binds to the nodal-related 2 (cyclops)andsox32 loci, accounting for the Nodal-independent expression of these genes in the yolk. Thus, Mxtx2 is anew actor in zebrafish mesendoderm induction, with a role upstreamof Nodal-related genes.doi:10.1016/j.ydbio.2010.05.084Program/Abstract # 52Roles of cilia in Gli protein activationAimin Liu, Huiqing Zeng, Amber Hoover, Jinping JiaDept. of Biol., The Penn State Univ., University Park, PA, USAHedgehog (Hh) family of signaling molecules plays pivotal roles incell proliferation and cell fate determination during developmentthrough regulating the activities of the Gli-family proteins. Ourprevious genetic studies indicated that cilia are required forintracellular transduction of Hh signals to Gli proteins in developingmouse embryos. In the current study, we are investigating themolecular mechanisms underlying the roles of cilia in regulating Gliactivities. One unresolved issue in this field has been the roles of ciliain Gli1 activation. Previous studies showed that over-expression ofGli1 in cultured cells activates Hh signaling in the absence of cilia;however, the requirement for cilia in Gli1 activation in vivo has beendifficult to address due to diminished Gli1 transcription in ciliamutants. In the current study, we restore Gli1 transcription in ciliamutants through a knock-in approach and find that Gli1 protein failsto activate Hh pathway in vivo in the absence of cilia, suggesting thatGli1 activation in vivo requires cilia. Subsequently, we show that allthree mouse Gli-family members share the same molecular mechan-ism for their ciliary localization, which appears to be independent oftheir phosphorylation and ubiquitylation. Finally, we show thatSuppressor of Fused (Sufu), an essential negative regulator of Hhsignaling, exerts its repressive function on Gli proteins independentof cilia. Association between Sufu and Gli proteins is independent ofcilia and is critical for proper translocation of both proteins.doi:10.1016/j.ydbio.2010.05.085Program/Abstract # 53The signaling properties in MAPK cycle are modulated by itsdownstream substratesTsuyoshi Hirashima