The ATP-binding cassette (ABC) transporter multidrug resistance protein 1 (MRP1/ABCC1) is responsible for the cellular export of a chemically diverse array of xenobiotics and endogenous compounds. Arsenic, a human carcinogen, is a high-affinity MRP1 substrate as arsenic triglutathione [As(GS) 3 ]. In this study, marked differences in As(GS) 3 transport kinetics were observed between MRP1-enriched membrane vesicles prepared from human embryonic kidney 293 (HEK) (K m 3.8 mM and V max 307 pmol/mg per minute) and HeLa (K m 0.32 mM and V max 42 pmol/mg per minute) cells. Mutant MRP1 lacking N-linked glycosylation [Asn19/23/1006Gln; sugar-free (SF)-MRP1] expressed in either HEK293 or HeLa cells had low K m and V max values for As(GS) 3 , similar to HeLa wild-type (WT) MRP1. When prepared in the presence of phosphatase inhibitors, both WT-and SF-MRP1-enriched membrane vesicles had a high K m value for As(GS) 3 (3-6 mM), regardless of the cell line. Kinetic parameters of As(GS) 3 for HEKAsn19/23Gln-MRP1 were similar to those of HeLa/HEK-SF-MRP1and HeLa-WT-MRP1, whereas those of single glycosylation mutants were like those of HEK-WT-MRP1. Mutation of 19 potential MRP1 phosphorylation sites revealed that HEK-Tyr920Phe/ Ser921Ala-MRP1 transported As(GS) 3 like HeLa-WT-MRP1, whereas individual HEK-Tyr920Phe-and -Ser921Ala-MRP1 mutants were similar to HEK-WT-MRP1. Together, these results suggest that Asn19/Asn23 glycosylation and Tyr920/Ser921 phosphorylation are responsible for altering the kinetics of MRP1-mediated As(GS) 3 transport. The kinetics of As(GS) 3 transport by HEK-Asn19/23Gln/Tyr920Glu/Ser921Glu were similar to HEK-WT-MRP1, indicating that the phosphorylation-mimicking substitutions abrogated the influence of Asn19/23Gln glycosylation. Overall, these data suggest that cross-talk between MRP1 glycosylation and phosphorylation occurs and that phosphorylation of Tyr920 and Ser921 can switch MRP1 to a lower-affinity, higher-capacity As(GS) 3 transporter, allowing arsenic detoxification over a broad concentration range.

Early HIV infection causes substantial mucosal and systemic immune activation, and gut CD4 T-cell dysfunction. One year of ART improved but did not normalize most parameters, regardless of intensification with raltegravir and maraviroc, and did not restore mucosal Th17 function.

These studies will evaluate the ability of probiotics as a safe and tolerable therapeutic intervention to reduce systemic immune activation and to accelerate gut immune restoration in people living with HIV.