Identification, characterization, and function analysis of the VIT family in Phaeodactylum tricornutum

Iron is an essential microelement for all living organisms. The vacuolar iron transporters (VIT) gene family is found in various species, including yeast, fungi, protozoa, and plants, where it plays a crucial role in sequestration, homeostasis, and tolerance of the heavy metals, particularly iron and manganese. However, the presence and function of VIT genes in marine phytoplankton have not been previously reported. The study aims to identify the VIT family within the marine diatom Phaeodactylum tricornutum and to analyze the function of these genes. We conducted a comprehensive analysis of the VIT genes in P. tricornutum genome, examining their phylogenetic relationship, physicochemical properties, gene structures, conserved motifs, domains, expression profile, and cis-acting elements using in silico methods. Function analysis were performed through complementation experiments and the expression of eGFP fusion protein in yeast. Four members of the VIT family were identified in P. tricornutum. All belonging to the VTL (VIT like) group in phylogenetic tree and containing a VIT1 domain. These genes are distributed across chromosomes 2, 4, and 13, with tandem duplication of the PtVTL1 and PtVTL2 contributed to the expansion of this gene family. Expression profile showed that the PtVTL3 is induced to express highly under light condition, others are induced to express highly under dark. PtVTL2 is highly induced to express at low Fe condition, and PtVTL3 is highly induced to express at high Fe condition. Analysis of cis-acting regulatory elements indicated that these genes are primarily involved in responses to environmental stress and phytohormones. Heterologous expression of PtVTL3 successfully rescued the iron-sensitive phenotype in yeast mutant △ccc1. The expression of eGFP-PtVTL3 fusion protein in yeast demonstrated that PtVTL3 is located to the tonoplast. These findings suggest that PtVTL3 function to transport Fe²⁺ across the tonoplast into the vacuole, thereby maintaining iron homeostasis in yeast. Four PtVTL genes were identified in the genome of P. tricornutum, with PtVTL3 playing a key role in iron transport at the tonoplast, highlighting its potential significance in iron homeostasis in marine diatoms.