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Two isoforms of the A subunit of the vacuolar H+-ATPase in Lycopersicon esculentum: highly similar proteins but divergent patterns of tissue localization

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Abstract

The plant vacuolar H+-translocating ATPase (V-ATPase, EC 3.6.1.34) generates a H+ electro-chemical gradient across the tonoplast membrane. We isolated two full-length cDNA clones (VHA-A1 and VHA-A2) from tomato (Lycopersicon esculentum Mill. cv. Large Cherry Red) coding for two isoforms of the V-ATPase catalytic subunit (V-ATPases A1 and A2). The cDNA clones encoding the two isoforms share 90% identity at the nucleotide level and 96% identity at the amino acid level. The 5′- and 3′-untranslated regions, however, are highly diverse. Both V-ATPase A1 and A2 isoforms encode polypeptides of 623 amino acids, with calculated molecular masses of 68,570 and 68,715, respectively. The expression of VHA-A1 and accumulation of V-ATPase A1 polypeptide were ubiquitous in all tissues examined. In response to salinity, the abundances of both transcript (VHA-A1) and protein (V-ATPase A1) of the A1 isoform in leaves were nearly doubled. In contrast to the A1 isoform, VHA-A2 transcript and V-ATPase A2 polypeptide were only detected in abundance in roots, and in minor quantities in mature fruit. In roots, accumulation of transcripts and polypeptides did not change in response to salinity for either isoform. Subcellular localization indicated that the highest levels of both V-ATPase A1 and A2 isoforms were in the tonoplast. However, significant quantities of both isoforms were detected in membranes associated with endoplasmic reticulum and/or Golgi. Immunoprecipitation of dissociated V1 domains using isoform-specific antibodies showed that V1 domains consist of either V-ATPase A1 or A2 catalytic subunit isoforms.

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Abbreviations

ER :

Endoplasmic reticulum

EST :

Expressed sequence tag

IDPase :

Inosine diphosphatase

V-ATPase :

Vacuolar H+-translocating ATPase

V-PPase :

Vacuolar H+-pyrophosphatase

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Acknowledgements

We thank Prof. R. Ratajczak (Darmstadt University of Technology, Darmstadt, Germany) for antibodies (ATP95) for the detection of V-ATPase holoenzyme, Prof. P.A. Rea (University of Pennsylvania, PA, USA) for antibodies (326) for the detection of vacuolar H+-pyrophosphatase and Dr. K.D. Hirschi (Baylor College of Medicine, Houston, TX) for providing BiP and PAQ1 antibodies. This work was supported in part by the U.S. Department of Agriculture (NRICGP #98-35100-6125 to M.L.B.) and the Texas Agricultural Experiment Station. The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. section 1734 solely to indicate this fact.

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Authors and Affiliations

  1. Institute of Plant Genomics and Biotechnology/Department of Horticulture, Texas A & M University, College Station, TX 77843, USA

    Umesh K. Bageshwar, Suparna Taneja-Bageshwar & Marla L. Binzel

  2. Department of Plant Biology and Pathology, Rutgers University, 59 Dudley Road, New Brunswick, NJ 08901, USA

    Hisham M. Moharram

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  1. Umesh K. Bageshwar
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  2. Suparna Taneja-Bageshwar
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  3. Hisham M. Moharram
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  4. Marla L. Binzel
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Corresponding author

Correspondence to Marla L. Binzel.

Additional information

The nucleotide sequences for the V-ATPase A1 and V-ATPase A2 cDNA clones have been deposited in the GenBank database under the GenBank Accession Numbers AY177247 and AY178911, respectively.

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Bageshwar, U.K., Taneja-Bageshwar, S., Moharram, H.M. et al. Two isoforms of the A subunit of the vacuolar H+-ATPase in Lycopersicon esculentum: highly similar proteins but divergent patterns of tissue localization. Planta 220, 632–643 (2005). https://doi.org/10.1007/s00425-004-1377-4

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