H2O2 and H2O transport by Medicago truncatula PIP2 aquaporins

CHEVRIAU, JONATHAN; ZERBETTO DE PALMA, GERARDO; CANESSA FORTUNA, AGUSTINA; VITALI, VICTORIA; ZEIDA, ARI; ALLEVA, KARINA

Rosario

Congreso; L Reunión Anual de la Sociedad Argentina de Biofísica; 2022

Several MIP (membrane intrinsic protein) channels transport H2O and hydrogen peroxide(H2O2) through the cell membrane. H2O2 transport occurs in different evolutive groups,and there is evidence that supports that this transport is an ancestral character of thefamily as well as the H2O transport. We are interested in understanding the differencesbetween H2O and H2O2 transport in isoforms of the plant PIP (plasma membrane intrinsicprotein) subfamily, due to their relevance in plant physiology.In previous studies, we showed that two PIPs from M. truncatula (MtPIP2;2 and MtPIP2;3)are efficient H2O and H2O2 channels. Now, our goal is to explore the particularities of thecombined H2O2 and H 2O transport by both in vivo and in silico approaches. Wecharacterized H2O and H2O2 permeation events by long unbiased molecular dynamicssimulations, finding that both molecules can cross the pore in a mixed single file, but withdifferent characteristic interactions throughout the pore. Further, there are densitieschanges between open and closed states of the pore, suggesting that the closed statecould be more restrictive towards H2O2 than to H2O. We also evaluated the residencetimes for H2O2, finding that residence times in the open pore are higher around theselectivity filter zone, while in the closed pore, the higher residence times were observedin the cytoplasmic constriction area.We overexpressed both MtPIP2;3 and MtPIP2;2 in X. laevis oocytes and performed swellingassays in the presence of different concentrations of external H2O2, both for the open andclosed states of these channels. Our preliminary results suggest that H2O2 did not altersignificantly water transport rates for any of the two MtPIP2. These results suggest thatboth molecules can be transported in a coordinated way, carrying out differentinteractions inside the pore. Further experiments and simulations will allow us to discloseif these distinct mechanisms are physiologically relevant