Project description:The aim of this study is to understand the mechanisms of TDP-43 neurotoxicity. Here, we perform a RNA-Seq analysis in TDP-43 gain-of-fucntion (GOF) , TDP-43 loss-of-function and wild-type late pupae heads (73-90 hours APF) and perform TDP-43 GOF vs wild type and TDP-43 LOF vs wild-type differential expression analysis to show that both mechanisms presents defects in ecdysone receptor (ECR)-dependeint transcriptional program switching, and strongly deregulate expression from the neuronal microtubule associated protien Map205. RNA-seq was performed in two wild-type D.melanogaster biological replicates (Canton S, w1118 ), four biological replicates for TDP-43 (LOF) with two distinct genotypes (dTDP-43Δ142/Df(2R)106,dTDP-43Δ23/Δ142 ) and two TDP-43 GOF biological replicates (act5c>dTDP-43 ).
Project description:To understand how the NAC transcription factor KIL1 regulates age-induced senescence and cell death in maize silks, we need to get a genome-wide view on its downstream targets. We propose to compare the transcriptome profiles of GOF and LOF transgenic silk tissue with the profile of wild-type B104 silk. 1 cm of basal part of silk from rings 6-10 from plants harboring the dominant-negative repressor proSILK1:KIL1-SRDX, proSILK1:KIL1 overexpressing line, and wild type B104 will be harvested at 11 DASE. This will allow to compare and contrast the expression profiles of KIL1 LOF and GOF mutants with transcriptome data derived from wild type senescent silk.
Project description:NOTCH1 is a transmembrane receptor that initiates the Notch signaling pathway involved in embryonic development and maintenance of adult tissue homeostasis. The extracellular part of NOTCH1 is composed largely of EGF-like domains (EGFs) many of which can be O-fucosylated by protein O-fucosyltransferase 1 (POFUT1). O-fucosylation of NOTCH1 is necessary for its function. The Notch pathway is deregulated in many cancers, and alteration of POFUT1 has been reported in some cancers. Using the Biomuta and COSMIC databases, we selected 9 NOTCH1 variants that could cause a change in O-fucosylation of key EGFs. Cell-based N1 signaling assays, Notch ligand-binding assays and cell surface N1 analysis were used to determine the effect of each mutation on Notch activation. Then mass spectral glycoproteomic site mapping was used to identify alterations in the O-fucosylation of EGFs containing the selected mutation. One of the variants had few effects, two lead to a gain of function (GOF) and six to a loss of function (LOF). Most GOF and LOF could be associated with a change in O-fucosylation. Finally, by comparing our results with known NOTCH1 alterations in cancers from which our mutations originated, we were able to establish a correlation between our results and the known GOF or LOF of NOTCH1 in these cancers. This study shows that point mutations in NOTCH1 can lead to alterations in O-fucosylation that deregulate the Notch pathway and be associated with cancer processes.