Abstract
Plant metal tolerance proteins (MTPs) comprise a family of membrane divalent cation transporters that play essential roles in plant mineral nutrition maintenance and heavy metal stresses resistance. However, the evolutionary relationships and biological functions of MTP family in tobacco remain unclear. In the present study, 26, 13, and 12 MTPs in three main Nicotiana species (N. tabacum, N. sylvestris, and N. tomentosiformis) were identified and designated, respectively. The phylogenetic relationships, gene structures, chromosome distributions, conserved motifs, and domains of NtMTPs were systematic analyzed. According to the phylogenetic features, 26 NtMTPs were classified into three major substrate-specific groups that were Zn-cation diffusion facilitators (CDFs), Zn/Fe-CDFs, and Mn-CDFs, and seven primary groups (1, 5, 6, 7, 8, 9, and 12). All of the NtMTPs contained a modified signature sequence and the cation_efflux domain, whereas some of them also harbored the ZT_dimer. Evolutionary analysis showed that NtMTP family of N. tabacum originated from its parental genome of N. sylvestris and N. tomentosiformis, and further underwent gene loss and expanded via one segmental duplication event. Moreover, the prediction of cis-acting elements (CREs) and the microRNA target sites of NtMTP genes suggested the diverse and complex regulatory mechanisms that control NtMTPs gene expression. Expression profile analysis derived from transcriptome data and quantitative real-time reverse transcription-PCR (qRT-PCR) analysis showed that the tissue expression patterns of NtMTPs in the same group were similar but varied among groups. Besides, under heavy metal toxicity, NtMTP genes exhibited various responses in either tobacco leaves or roots. 19 and 15 NtMTPs were found to response to at least one metal ion treatment in leaves and roots, respectively. In addition, NtMTP8.1, NtMTP8.4, and NtMTP11.1 exhibited Mn transport abilities in yeast cells. These results provided a perspective on the evolution of MTP genes in tobacco and were helpful for further functional characterization of NtMTP genes.