A differential subcellular localization of two copper transporters from the COPT family suggests distinct roles in copper homeostasis in Physcomitrium patens

The moss Physcomitrium (Physcomitrella) patens is a bryophyte that provides genetic information about the adaptation to the life on land by early Embryophytes and is a reference organism for comparative evolutionary studies in plants. Copper is an essential micronutrient for every living organism, its transport across the plasma membrane is achieved by the copper transport protein family COPT/CTR. Two genes related to the COPT family were identified in Physcomitrella patens, PpaCOPT1 and PpaCOPT2. Homology modelling of both proteins showed the presence of three putative transmembrane domains (TMD) and the Mx3M motif, constituting a potential Cu + selectivity filter present in other members of this family. Functional characterization of PpaCOPT1 and PpaCOPT2 in the yeast mutant ctr1Δctr3Δ restored its growth on medium with non-fermentable carbon sources at micromolar Cu concentrations, providing support that these two moss proteins function as high affinity Cu + transporters. Localization of PpaCOPT1 and PpaCOPT2 in yeast cells was observed at the tonoplast and plasma membrane, respectively. The heterologous expression of PpaCOPT2 in tobacco epidermal cells co-localized with the plasma membrane marker. Finally, only PpaCOPT1 was expressed in seven-day old protonema and was influenced by extracellular copper levels. This evidence suggests different roles of PpaCOPT1 and PpaCOPT2 in copper homeostasis in Physcomitrella patens.