GH3 Gene Family Identification in Chinese White Pear (Pyrus bretschneideri) and the Functional Analysis of PbrGH3.5 in Fe Deficiency Responses in Tomato

Iron (Fe) deficiency poses a major threat to pear (<i>Pyrus</i> spp.) fruit yield and quality. The <i>Gretchen Hagen 3</i> (<i>GH3</i>) plays a vital part in plant stress responses. However, the <i>GH3</i> gene family is yet to be characterized, and little focus has been given to the function of the <i>GH3</i> gene in Fe deficiency responses. Here, we identified 15 GH3 proteins from the proteome of Chinese white pear (<i>Pyrus bretschneideri</i>) and analyzed their features using bioinformatics approaches. Structure domain and motif analyses showed that these PbrGH3s were relatively conserved, and phylogenetic investigation displayed that they were clustered into two groups (GH3 I and GH3 II). Meanwhile, <i>cis</i>-acting regulatory element searches of the corresponding promoters revealed that these <i>PbrGH3s</i> might be involved in ABA- and drought-mediated responses. Moreover, the analysis of gene expression patterns exhibited that most of the <i>PbrGH3s</i> were highly expressed in the calyxes, ovaries, and stems of pear plants, and some genes were significantly differentially expressed in normal and Fe-deficient pear leaves, especially for <i>PbrGH3.5</i>. Subsequently, the sequence of <i>PbrGH3.5</i> was isolated from the pear, and the transgenic tomato plants with <i>PbrGH3.5</i> overexpression (OE) were generated to investigate its role in Fe deficiency responses. It was found that the OE plants were more sensitive to Fe deficiency stress. Compared with wild-type (WT) plants, the rhizosphere acidification and ferric reductase activities were markedly weakened, and the capacity to scavenge reactive oxygen species was prominently impaired in OE plants under Fe starvation conditions. Moreover, the expressions of Fe-acquisition-associated genes, such as <i>SlAHA4</i>, <i>SlFRO1</i>, <i>SlIRT1</i>, and <i>SlFER</i>, were all greatly repressed in OE leaves under Fe depravation stress, and the free IAA level was dramatically reduced, while the conjugated IAA contents were notably escalated. Combined, our findings suggest that pear <i>PbrGH3.5</i> negatively regulates Fe deficiency responses in tomato plants, and might help enrich the molecular basis of Fe deficiency responses in woody plants.