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A plant plasma-membrane H⁺-ATPase promotes yeast TORC1 activation via its carboxy-terminal tail.

ABSTRACT: The Target of Rapamycin Complex 1 (TORC1) involved in coordination of cell growth and metabolism is highly conserved among eukaryotes. Yet the signals and mechanisms controlling its activity differ among taxa, according to their biological specificities. A common feature of fungal and plant cells, distinguishing them from animal cells, is that their plasma membrane contains a highly abundant H⁺-ATPase which establishes an electrochemical H⁺ gradient driving active nutrient transport. We have previously reported that in yeast, nutrient-uptake-coupled H⁺ influx elicits transient TORC1 activation and that the plasma-membrane H⁺-ATPase Pma1 plays an important role in this activation, involving more than just establishment of the H⁺ gradient. We show here that the PMA2 H⁺-ATPase from the plant Nicotiana plumbaginifolia can substitute for Pma1 in yeast, to promote H⁺-elicited TORC1 activation. This H⁺-ATPase is highly similar to Pma1 but has a longer carboxy-terminal tail binding 14-3-3 proteins. We report that a C-terminally truncated PMA2, which remains fully active, fails to promote H⁺-elicited TORC1 activation. Activation is also impaired when binding of PMA2 to 14-3-3 s is hindered. Our results show that at least some plant plasma-membrane H⁺-ATPases share with yeast Pma1 the ability to promote TORC1 activation in yeast upon H⁺-coupled nutrient uptake.

SUBMITTER: Saliba E

PROVIDER: S-EPMC7910539 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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A plant plasma-membrane H+-ATPase promotes yeast TORC1 activation via its carboxy-terminal tail.

Saliba Elie E Primo Cecilia C Guarini Nadia N André Bruno B

Scientific reports 20210226 1

The Target of Rapamycin Complex 1 (TORC1) involved in coordination of cell growth and metabolism is highly conserved among eukaryotes. Yet the signals and mechanisms controlling its activity differ among taxa, according to their biological specificities. A common feature of fungal and plant cells, distinguishing them from animal cells, is that their plasma membrane contains a highly abundant H+-ATPase which establishes an electrochemical H+ gradient driving active nutrient ...[more]

PMID: 33637787

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Project description:The crystal structures of the Ca(2+)- and H(+)-ATPases shed light into the membrane embedded domains involved in binding and ion translocation. Consistent with site-directed mutagenesis, these structures provided additional evidence that membrane-spanning segments M4, M5, M6 and M8 are the core through which cations are pumped. In the present study, we have used alanine/serine scanning mutagenesis to study the structure-function relationships within M6 (Leu-721-Pro-742) of the yeast plasma membrane ATPase. Of the 22 mutants expressed and analyzed in secretory vesicles, alanine substitutions at two well conserved residues (Asp-730 and Asp-739) led to a complete block in biogenesis; in the mammalian P-ATPases, residues corresponding to Asp-730 are part of the cation-binding domain. Two other mutants (V723A and I736A) displayed a dramatic 20-fold increase in the IC(50) for inorganic orthovanadate compared to the wild-type control, accompanied by a significant reduction in the K(m) for Mg-ATP, and an alkaline shift in the pH optimum for ATP hydrolysis. This behavior is apparently due to a shift in equilibrium from the E(2) conformation of the ATPase towards the E(1) conformation. By contrast, the most striking mutants lying toward the extracellular side in a helical structure (L721A, I722A, F724A, I725A, I727A and F728A) were expressed in secretory vesicles but had a severe reduction of ATPase activity. Moreover, all of these mutants but one (F728A) were unable to support yeast growth when the wild-type chromosomal PMA1 gene was replaced by the mutant allele. Surprisingly, in contrast to M8 where mutations S800A and E803Q (Guerra et al., Biochim. Biophys. Acta 1768: 2383-2392, 2007) led to a dramatic increase in the apparent stoichiometry of H(+) transport, three substitutions (A726S, A732S and T733A) in M6 showed a reduction in the apparent coupling ratio. Taken together, these results suggest that M6 residues play an important role in protein stability and function, and probably are responsible for cation binding and stoichiometry of ion transport as suggested by homology modeling.

Dataset Information

A plant plasma-membrane H⁺-ATPase promotes yeast TORC1 activation via its carboxy-terminal tail.

Publications

A plant plasma-membrane H<sup>+</sup>-ATPase promotes yeast TORC1 activation via its carboxy-terminal tail.

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