Dissection of TMEM165 function in Golgi glycosylation and its Mn2+ sensitivity

Abstract

Since 2012, the interest for TMEM165 increased due to its implication in a rare genetic human disease named TMEM165-CDG (Congenital Disorder(s) of Glycosylation). TMEM165 is a Golgi localized protein, highly conserved through evolution and belonging to the uncharacterized protein family 0016 (UPF0016). Although the precise function of TMEM165 in glycosylation is still controversial, our results highly suggest that TMEM165 would act as a Golgi Ca2+/Mn2+ transporter regulating both Ca2+ and Mn2+ Golgi homeostasis, the latter is required as a major cofactor of many Golgi glycosylation enzymes. Strikingly, we recently demonstrated that besides its role in regulating Golgi Mn2+ homeostasis and consequently Golgi glycosylation, TMEM165 is sensitive to high manganese exposure. Members of the UPF0016 family contain two particularly highly conserved consensus motifs E-φ-G-D-[KR]-[TS] predicted to be involved in the ion transport function of UPF0016 members. We investigate the contribution of these two specific motifs in the function of TMEM165 in Golgi glycosylation and in its Mn2+ sensitivity. Our results show the crucial importance of these two conserved motifs and underline the contribution of some specific amino acids in both Golgi glycosylation and Mn2+ sensitivity.

From the Uncharacterized Protein Family 0016 to the GDT1 family: Molecular insights into a newly-characterized family of cation secondary transporters

“…One of the possible mechanisms of transport with these three actors consists of a Ca 2+ -Mn 2+ /H + antiporter, where Ca 2+ and Mn 2+ would be transported at the Golgi level in one direction in exchange of protons. However, and in accordance with the limited and relatively indirect evidence of transport of protons, it was suggested that these UPF0016 members would work as Ca 2+ /Mn 2+ antiporters [ 23 , 29 ]. In A. thaliana , data from different studies led to the conclusion that Mn 2+ cations and, if transported, Ca 2+ cations would also be transported in the same direction, towards the chloroplast lumen [ 3 – 7 ], with indication of transport of protons only for PAM71 [ 3 ].…”

“…Nevertheless, based on data interpretation, the human and yeast orthologs TMEM165 and Gdt1p could transport Ca 2+ and Mn 2+ from the cytosol to the Golgi lumen in exchange of protons from the Golgi to the cytosol, if working as Ca 2+ -Mn 2+ /H + antiporters. In case of a Ca 2+ /Mn 2+ antiporter, data suggest that Mn 2+ would be imported in the Golgi in exchange of Ca 2+ [ 23 , 29 ]. In plants, Mn 2+ and, if transported, Ca 2+ would be transported from the cytosol to the stroma via CMT1, and then from the stroma to the thylakoid lumen via PAM71 [ 3 – 7 ].…”

“…Altogether, this suggests that these residues are essential for the Ca 2+ transport activity of Gdt1p (given that these mutated proteins are produced and well localized as verified by [ 59 ]). In human, the ability of wild-type or mutated TMEM165 to rescue glycosylation defects observed in TMEM165-KO cells was similarly used to investigate the involvement of the residues of the conserved motifs, as well as of flanking residues, in protein functionality [ 29 ]. Interestingly, the mutated proteins from the motifs 248 Glu, 111 Asp & 251 Asp, 113 Thr, and 253 Ser, as well as from the flanking residue 114 Phe, were all unable to restore glycosylation.…”

The Mechanism of Zinc Sulfate in Improving Fertility in Obese Rats Analyzed by Sperm Proteomic Analysis

“…However, data obtained in recent study give several indications of their implication in calcium and manganese homeostasis [47]. TM165 supplies Ca2+ and Mn2+ to the Golgi complex in exchange for H+ to sustain the functions of lactose synthase and potentially other glycosyltransferases [48,49]. The human Golgi protein TM165 can transport calcium and manganese in yeast and bacterial cells [50].…”

Novel role for the Golgi membrane protein TMEM165 in control of migration and invasion for breast carcinoma

“…Possible explanations include that TMEM165, a Golgi transmembrane protein and putative ion transporter, is acting as a Mn2+/Ca2+ transporter moving Mn2+ from the Golgi lumen to the cytosol. Evidence supports a role for TMEM165 in Mn++ transport as the TMEM165 protein is degraded by high cytosolic Mn2+ concentration [14,40]. In breast cancer, calcium ATPases are altered in expression levels and activity and these findings would further complicate the control of ion homeostasis [41,42].…”