Relevance of 3´Unstranlated region (3´UTR) in regulation of mRNA beta1 integrin (Itgb1) expression

NAIPAUER J.;; GATTELLI A.; LAMARRE J.; COSO O; KORDON E.

Villa Carlos Paz, Córdoba, Argentina.

Congreso; XLIV Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular,; 2008

SAIB

Integrins belong to a superfamily of cell surface adhesion receptors that play a relevant role in mammary gland biology. Specifically, in this tissue, it has been demonstrated that b1-integrin plays a critical role during normal development and tumor formation. Therefore, analyzing expression regulation of this gene may facilitate the understanding of both processes. Based on numerous studies that have implicated the 3’UTR sequences in the regulation of mRNA levels, we have examined gene products of b1 integrin in normal and neoplastic mammary glands by 3’RACE-PCR. By cloning and sequencing these reaction products, we have found a new b1-integrin mRNA species, 578bp shorter than the previously reported one. By sequence analysis we have determined that this different species arise from the use of alternative polyadenylation sites. Our analysis also revealed the presence of two AU-rich element (ARE) sequences, localized between those sites and, therefore, absent in the shorter mRNA species. By semiquantitative RT-PCR and RNAse protection assays, we determined that both forms are present in normal and neoplastic tissue. We have found that the previously reported longer mRNA is more abundant, although each stage of mammary gland development correlates with a specific ratio between the longer and the shorter form. To perform a more detailed analysis, we have sub-cloned the two different 3’UTR sequences into reporter vectors downstream of a CMV driven luciferase gene. Then, mRNA stability assays were performed using the TET-OFF system. The results indicated that the longer form is less stable, suggesting that the ARE sequences may play a relevant role on this mRNA stability. Therefore, the 3’UTR regulating sequences may provide another regulatory instance to determine more precisely the b1-integrin abundance in different stages of mammary gland development.