Naslov
Epidermal growth factor stimulates translocation of the class II phosphoinositide 3-kinase C2beta to the nucleus

Autori
Banfić, Hrvoje ; Mise, N ; Gu, Y ; Domin, Jan

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, neobjavljeni rad, znanstveni

Skup
Gordon Research Conference "Signal Transduction within the Nucleus"

Mjesto i datum
Ventura (CA), Sjedinjene Američke Države, 25.03.2007. - 30.03.2007

Vrsta sudjelovanja
Poster

Vrsta recenzije
Međunarodna recenzija

Ključne riječi
PI3KC2beta; EGF; nucleus

Sažetak
In native cells calpain-mediated activation of phosphoinositide 3-kinase C2 (PI3K-C2  was observed in nuclei following partial hepatectomy and during the G2/M phase of cell cycle. Differentiation of HL-60 cells with all-trans retinoic acid increased PI3K-C2 enzyme activity in nuclei by a mechanism that involves tyrosine phosphorylation. In this study we have used HEK293 cells to visualise nuclear compartmentalization of PI3K-C2 and examine the possibility of its translocation to the nucleus following growth factor stimulation. Membrane-depleted nuclei were isolated by osmotic shock and low speed centrifugation using sucrose cushion in the presence of 1% Triton and 0.1% deoxycholate. This enriched preparation of nuclei was free of  -tubulin and contained less than 5% of the total cellular content of PI3K-C2 enzyme. When nuclear matrix and chromatin were isolated from membrane-depleted nuclei and subjected to Western blot analysis, PI3K-C2 was localized in the nuclear matrix. Furthermore, confocal analysis of membrane depleted nuclei revealed that the enzyme co-localized with lamin A/C supporting its localization in the nuclear matrix. Stimulation of HEK293 cells with epidermal growth factor (EGF) increased levels of PI3K-C2 in nuclei in a time dependent manner becoming maximal by 90 minutes. Transfection of HEK293 cells with C-terminal PI3K-C2 deletion mutants demonstrated that constructs lacking the C-terminal C2 domain were absent in membrane-depleted nuclei and EGF could not stimulate their translocation. Expression of the C-terminal PI3K-C2  C2 domain as an EGFP fusion protein revealed its presence in nuclei increased following EGF stimulation. In summary we have shown that the class II PI3K-C2 is: a) localized in the nuclear matrix, b) translocates into nuclei following stimulation with EGF and c) the translocation of PI3K-C2 to the nucleus is dependent upon the C-terminal C2 domain.

Izvorni jezik
Engleski

Znanstvena područja
Temeljne medicinske znanosti

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