TRANSCRIPTIONAL AND POST-TRANSLATIONAL ERK 1/2 REGULATION OF MKP-3 SPLICE VARIANTS IN BREAST CANCER CELLS

SILVANA IRIS NUDLER; ANA FERNANDA CASTILLO; JUAN MANUEL COHEN SABBAN; MARÍA MERCEDES MORI SEQUEIROS GARCÍA; CRISTINA PAZ

Mar del Plata

Congreso; LXI REUNIÓN ANUAL DE LA SOCIEDAD ARGENTINA DE INVESTIGACIÓN CLÍNICA (SAIC); 2016

Sociedad Argentina de Investigación Clínica (SAIC)

MKP-3 is a member of the family of mitogen-activated protein kinase phosphatase (MKPs) that specifically dephosphorylates ERK 1/2, acting as a key effector of the MAPK signal pathway. Variable expression of MKP-3 was detected in different cancer types. In human tissues and cell lines, two splice variants of MKP-3 are expressed: the full transcript or isoform L, and isoform S, where exon 2 is skipped. The function of MKP-3 and its isoforms in breast cancer is still not well known. The aim of this work was to analyze and to compare the role of P-ERK on mRNA and protein stability of both isoforms in the breast cancer cell line MDA-MB-231. Real time RT-PCR analysis showed that both isoforms, S and L, are induced by a mitogenic stimuli (fetal bovine serum, FBS) in a time and ERK-dependent manner. The maximal induction of L and S isoforms was detected after 3 h of stimulation (1.4 and 1.8-fold, for L and S respectively). In cells stimulated with FBS for 3 h, a MEK (MAPK kinase) inhibitor (PD 98059) reduced L and S mRNA levels in a time dependent manner, reaching a 10 % of the initial value after 3 h of treatment. In actinomicyn D-treated cells, the decay of both isoforms was different. Indeed, mRNA of the L isoform showed higher stability as compared to the S isoform (half life: 60 vs. 38 min), while blockade of P-ERK signaling by PD 98059 reduced mRNA stability of both isoforms. Western blot analysis revealed the presence of L and S proteins in MDA-MB-231 cells, being L levels up-regulated by the MEK inhibitor. Together, our results show that P-ERK regulates the expression and stability of mRNAs of both isoforms of MKP-3 but only the stability of the L protein.