Identification and characterization of two genes encoding plasma membrane H+-ATPase in Cucumis sativus L.

Abstract

Two genes (CsHA2, CsHA3) homolog to the plasma membrane H ? -ATPase encoding sequences have been identified in cucumber cells. The full-length cDNA of these sequences comprise an open reading frame that encodes about 950 amino acid polypeptide with several potential transmembrane domains. Both sequences share a high homology with putative plasma membrane-associated proton pumps in higher plants, ranging from 84% with that of Arabidopsis thaliana to 90% with Sesbania rostrata. Phylogenetic analysis grouped the two isoforms into one group (subfamilies II) together with Kosteletzkya virginica KVATP1, Sesbania rostrata SRHA4, tobacco PMA4 and rice OSA7. The expression of CsHA isoforms at the mRNA level showed they different organ pattern. Transcript level of CsHA2 was detected in all vegetative organs, but more abundantly in leaves than in roots, while CsHA3 expression was limited only to the roots in immature as well as in mature plants. Moreover, the RT-PCR analysis of generative organs (flowers), fruits and dry seeds revealed differential expression pattern for CsHA2 and CsHA3 suggesting that these proteins could be involved in separate physiological processes.

The plasma membrane proton pump gene family in cucumber

“…These sequences comprised two genes (CsHA2 and CsHA3) identified by us earlier (Mlodzinska et al 2010). The phylogenetic analysis of CsHAs based on the protein alignment revealed the representatives of three (I, II and IV, Fig.…”

“…Likewise, two cucumber members of subfamily II, CsHA2 and CsHA4, also widely expressed in plant tissues, might encode the housekeeping proteins, despite the different expression levels in particular organs at the various developmental stages of cucumber (Fig. 3, Mlodzinska et al 2010). Such expression profiles suggest additionally that during plant development one CsHA gene can take over the role of another.…”

“…Obtained amplicons of CsHA genes were sequenced to confirm specificity of the PCR products and were deposited in the publicly available database (GenBank) as sequence tags (ESTs) ( Table 1). Specific primers necessary for CsHA2 and CsHA3 amplification were used according to Santi et al (2005) and Mlodzinska et al (2010). Finally, ten specific primer pairs were designed and used in experiments.…”

“…Using Blastn searches of cucumber whole genome shotgun reads from the GenBank database with 12 A. thaliana cDNAs encoding AtHA1-12 proteins as the query, we retrieved a total of ten sequences, which significantly matched with AtHAs. Two of them (CsHA2 and CsHA3) have been described earlier (Mlodzinska et al 2010). Next eight CsHA genes were named after the other according to similarity to earlier identified CsHA2 and CsHA3.…”

“…A higher level of the LHA4 transcript in darkness suggested its role in adaptation to light conditions (Mito et al 1996) while clear induction of LHA8 expression under salinity implied its importance in salt resistance (Kalampanayil and Wimmers 2001). Although HA genes in Arabidopsis, rice and Nicotiana have already been characterized, and recently, two cucumber HA genes (CsHA2 and CsHA3) have been identified (Mlodzinska et al 2010;Santi et al 2005), a systemic analysis of the HA gene family in cucumber has not been reported. The availability of the cucumber genome sequence (Huang et al 2009;Woycicki et al 2011) has provided an excellent opportunity for whole genome annotation, classification and comparative genomic research.…”