Identification and distinct regulation of three di/tripeptide transporters in Aspergillus oryzae

ABSTRACT The uptake of di/tripeptides is mediated by the proton-dependent oligopeptide transporter (POT) family. In this study, 3 POT family transporters, designated PotA, PotB, and PotC were identified in Aspergillus oryzae. Growth comparison of deletion mutants of these transporter genes suggested that PotB and PotC are responsible for di/tripeptide uptake. PotA, which had the highest sequence similarity to yeast POT (Ptr2), contributed little to the uptake. Nitrogen starvation induced potB and potC expression, but not potA expression. When 3 dipeptides were provided as nitrogen sources, the expression profiles of these genes were different. PrtR, a transcription factor that regulates proteolytic genes, was involved in regulation of potA and potB but not in potC expression. Only potC expression levels were dramatically reduced by disruption of ubrA, an orthologue of yeast ubiquitin ligase UBR1 responsible for PTR2 expression. Expression of individual POT genes is apparently controlled by different regulatory mechanisms.

Preferential dipeptide incorporation of Porphyromonas gingivalis mediated by proton-dependent oligopeptide transporter (Pot)

Multiple dipeptidyl-peptidases (DPPs) are present in the periplasmic space of Porphyromonas gingivalis, an asaccharolytic periodontopathic bacterium. Dipeptides produced by DPPs are presumed to be transported into the bacterial cells and metabolized to generate energy and cellular components. The present study aimed to identify a transporter responsible for dipeptide uptake in the bacterium. A real-time metabolic analysis demonstrated that P. gingivalis preferentially incorporated Gly-Xaa dipeptides, and then, single amino acids, tripeptides, and longer oligopeptides to lesser extents. Heterologous expression of the P. gingivalis serine/threonine transporter (SstT) (PGN_1460), oligopeptide transporter (Opt) (PGN_1518), and proton-dependent oligopeptide transporter (Pot) (PGN_0135) genes demonstrated that Escherichia coli expressing Pot exclusively incorporated Gly-Gly, while SstT managed Ser uptake and Opt was responsible for Gly-Gly-Gly uptake. Dipeptide uptake was significantly decreased in a P. gingivalis Δpot strain and further suppressed in a Δpot-Δopt double-deficient strain. In addition, the growth of the Δpot strain was markedly attenuated and the Δpot-Δopt strain scarcely grew, whereas the ΔsstT strain grew well almost like wild type. Consequently, these results demonstrate that predominant uptake of dipeptide in P. gingivalis is mostly managed by POT. We thus propose that Pot is a potential therapeutic target of periodontal disease and P. gingivalis-related systemic diseases.

Cloning, phylogenetic analysis, and postnatal expression of oligopeptide transporter PepT1 in gastrointestinal tract of kid goats receiving supplemental feed or pasture

This study aimed to clone the cDNA of PepT1, an H+-dependent oligopeptide transporter, from kid goats and examine effects of physiological development (suckling, weaning, and post-weaning) of the animal and feeding system (supplemental feeding vs. grazing) on peptide transport capability. A 2395 bp cDNA sequence of pept1 (GenBank: MH308024) was cloned and phylogenetic analysis revealed a high homology and structure similarity with PepT1 of sheep and cattle. The pept1 was expressed throughout the gastrointestinal tract of kid goats immediately after birth and during development. Relative abundance of pept1 decreased in all segments except the middle-jejunum during suckling, whereas its expression in most segments of small intestine increased with age after weaning and remained stable thereafter. Middle-jejunum was the predominant expression site and probably the main peptide absorption site. Supplemental feeding enhanced pept1 expression because it increased protein intake compared with grazing. No feeding system × age interaction was observed in most segments; the expression was age related during suckling and diet related during weaning and post-weaning, indicating that feeding system and age had independent effects on pept1 expression. These results indicate that PepT1 plays an important role for protein nutrition in neonatal goats, and its expression can be affected by feeding system.

In silico identification and expression analysis of Metal-nicotianamine transporter (YSL3) and Oligopeptide transporter 3 (OPT3) under Cd stress in Brassica oleracea var. acephala

Background: Metal-nicotianamine transporter (YSL) family protein belongs to the oligopeptide heavy metal transporter group, as characterized in Arabidopsis thaliana. Oligopeptide transporters (OPTs) are a group of membrane-localized proteins, involved in different transport mechanisms, contributing to nitrogen mobilization, glutathione transport and long-distance metal distribution. Metal-nicotianamine transporter gene 3 (YSL3) incorporates the oligopeptide transporter domain, found to transfer several heavy metals in diverse plant species, and among them cadmium transport in Brassica oleracea. Objective: To evaluate and confirm the expression of Metal-nicotianamine transporter (YSL3) under cadmium stress. Studied species: Brassica oleracea var. acephala Study site and dates: Brassica oleracea var. acephala samples were collected from Blagaj region, Bosnia and Herzegovina. Methods: Through a simple bioinformatic approach the interactome partner of Metal-nicotianamine transporter (YSL3) was discovered and annotated. Oligopeptide transporter 3 (OPT3) and Metal-nicotianamine transporter (YSL3) genes were checked for expression levels under cadmium stress. Results: We have identified a strong interacting partner of YSL3, later confirmed as Oligopeptide transporter 3 (OPT3) protein in Brassica oleracea. The in vitro expression analysis by using a qRT-PCR revealed a significant upregulation of YSL3 and OPT3, during Cd stress. Conclusions: These findings indicate that the represented in-silico approach, followed by in vitro gene expression study, successfully confirmed YSL3 and identified OPT3 as a new gene, in correlation to cadmium stress.