Abstracts Category : Other

Functional analysis of mutation in the linker domain of Smad3: Determination of nuclear localization and transcriptional activation potential of Smad3 mutant

by Matthildur Sigurbergsdttir Gurn

Slagaglpur s ea saglpur er sjkdmur sem getur auveldlega haft lfshttulegar afleiingar. Sjklingar finna yfirleitt ekki fyrir einkennum og uppgtvast glpurinn fyrir tilviljun, egar sin a lokum rofnar ea ef einstaklingar vera fyrir brri sarflysjun. sarglpur brjstholi er algengur msum bandvefssjkdmum bor vi Marfans og Ehler-Danlos heilkenni. Einnig virist sjkdmurinn ganga ttum ar sem stkkbreytingar msum genum hafa fundist fjlskyldum ar sem saglpur brjstholi virist arfgengur. slensk Erfagreining fann nlega eina slka stkkbreytingu slenskri fjlskyldu og er hn skou nnar essu verkefni. Stkkbreytingin er stasett tengisvi (linker domain) geninu Smad3 sem kar fyrir lykilprtein TGF boleiinni. Markmi verkefnisins var a framkalla stkkbreytinguna og framkvma henni virkniprf sem gefa betri skilning hrifum hennar. Til a greina hvort stkkbreytingin hafi hrif innflutning prteinsins kjarna var stkkbreytt Smad3 yfirtj frumum. Annars vegar var mtefnalitun framkvmd og greind lagsj. Hinsvegar voru framkvmdar tilraunir me luciferasa klguskju til a kanna umritunarvirkni stkkbreytta Smad3 prteinsins. Niurstur r essum tilraunum leiddu ljs a stkkbreytingin veldur v a Smad3 ferast ekki inn kjarnann lkt og villigerarprteini egar frumurnar voru rvaar me TGF. Einnig virtist stkkbreytta prteini ekki geta virkja umritun sama htt og villigerarprteini, bi rvuum og TGF rvuum frumum. : Aortic aneurysm is a condition in which the aorta bulges out in an abnormal way. Patients are most often asymptomatic until the aneurysm is diagnosed by accident or they end up suffering from aortic rupture or dissection which is potentially lethal. When the bulge forms in the chest portion of the aorta it is referred to as thoracic aortic aneursym (TAA). TAA is prevalent in some inherited connective tissue disorders such as Marfan syndrome and Ehler-Danlos syndrome and mutations in various genes have been found in families with history of aortic aneurysm. Recently, Decode Genetics found one such mutation in an Icelandic family. The mutation is located in the linker domain of Smad3 gene that encodes an essential protein in the TGF signaling pathway. The aim of this study was to generate that mutation and perform a functional analysis of the mutant. Overexpression, immunostaining and confocal imaging were used to determine subcellular localization of the protein and a luciferase reporter assay was performed to analyse the transcriptional activation of the mutated Smad3. Results revealed that the mutated Smad3 does not seem to be able to translocate to the nucleus compared to the wt Smad3 when cells are stimulated with TGF. The mutant also showed less transcriptional activation downstream of the TGF pathway compared to the wt Smad3 both in non-treated and TGF treated cells.