MKP-1 MODULATES ENDOPLASMIC RETICULUM STRESS EVENTS TRIGGERED BY CISPLATIN IN RENAL TUBULE CELLS

LUCIANA ANDREA CABRERA ESCOBAR; ANDREA BEATRIZ ACQUIER; CF MENDEZ; CRISTINA PAZ; ALEJANDRA BEATRIZ GOROSTIZAGA

Mar del Plata

Congreso; LXI Reunión anual de la SAIC; 2016

Sociedad Argentina de Investigación Clínica

1INBIOMED, UBA-CONICET, Departamento de Bioquimica Humana, Facultad de Medicina, Universidad de Buenos Aires. Argentina. 2Cátedra de Farmacologia, Facultad De Odontologia, Universidad de Buenos Aires. ArgentinaA wide variety of agents induce endoplasmic reticulum stress (RE) in different cell types. RE produces a complex signal transduction pathway known as Unfolding Protein Response that includes activation of the MAP kinases (MAPKs) ERK1/2, JNK1/2 and p38 which have different effects on survival or apoptosis. Stimuli that promote the activation of MAPKs induce the expression of different members of the family of MAPK phosphatases (MKPs), which promote the inactivation of these kinases. MKP-1 is a well characterized member of this family induced by several stimuli and able to dephosphorylate members of the three MAPKs subgroups. Cisplatin (CPT), a known chemotherapeutic agent, is widely used as RE inductor. The aim of this work was to determine if MAPK phosphatase 1 (MKP-1) modulates aspects related to RE in a human renal proximal tubule-derived cell line (HK-2) exposed to CPT. First, we analyzed MAPKs activation by CPT. Western blot analysis using specific antibodies against the phosphorylated forms of ERK1/2 showed that 50 mM CPT promotes ERK1/2 activation in a time-dependent manner. The increase was evident after 4 h of stimulation (6-fold) extending to 12 h. We also analyzed MKP-1 mRNA levels and we observed that CPT increases mRNA levels in a time- and concentration-dependent manner. In addition, we analyzed the expression of GRP78, a RE marker involved in cell survival by semiquantitative RT-PCR. We found that CPT increased GRP78 levels reaching a maximum at 6 h (3-fold). PD98059 (50 uM), an inhibitor of ERK1/2 activation, prevented the effect of CPT on GRP78 levels, while SB203580 and SP-600125 (JNK and p38 inhibitors, respectively) had no effect, thereby suggesting that GRP78 expression is modulated by ERK1/2. Moreover, in cells transfected with a construction for transient expression of flag-MKP-1 recombinant protein, MTT assay showed that transfection with MKP-1 modulates cell viability. Collectively, our data suggest that MKP-1 is induced by CPT and may contribute to turn off MAPKs-dependent events triggered by CPT.