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Assembly of B4GALT1/ST6GAL1 heteromers in the Golgi membranes involves lateral interactions via highly charged surface domains.

ABSTRACT: ?-1,4-Galactosyltransferase 1 (B4GALT1) and ST6 ?-galactoside ?-2,6-sialyltransferase 1 (ST6GAL1) catalyze the successive addition of terminal ?-1,4-linked galactose and ?-2,6-linked sialic acid to N-glycans. Their exclusive interaction in the Golgi compartment is a prerequisite for their full catalytic activity, whereas a lack of this interaction is associated with cancers and hypoxia. To date, no structural information exists that shows how glycosyltransferases functionally assemble with each other. Using molecular docking simulations to predict interaction surfaces, along with mutagenesis screens and high-throughput FRET analyses in live cells to validate these predictions, we show here that B4GALT1 and ST6GAL1 interact via highly charged noncatalytic surfaces, leaving the active sites exposed and accessible for donor and acceptor substrate binding. Moreover, we found that the assembly of ST6GAL1 homomers in the endoplasmic reticulum before ST6GAL1 activation in the Golgi utilizes the same noncatalytic surface, whereas B4GALT1 uses its active-site surface for assembly, which silences its catalytic activity. Last, we show that the homomeric and heteromeric B4GALT1/ST6GAL1 complexes can assemble laterally in the Golgi membranes without forming cross-cisternal contacts between enzyme molecules residing in the opposite membranes of each Golgi cisterna. Our results provide detailed mechanistic insights into the regulation of glycosyltransferase interactions, the transitions between B4GALT1 and ST6GAL1 homo- and heteromers in the Golgi, and cooperative B4GALT1/ST6GAL1 function in N-glycan synthesis.

SUBMITTER: Khoder-Agha F

PROVIDER: S-EPMC6768654 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Assembly of B4GALT1/ST6GAL1 heteromers in the Golgi membranes involves lateral interactions via highly charged surface domains.

Khoder-Agha Fawzi F Harrus Deborah D Brysbaert Guillaume G Lensink Marc F MF Harduin-Lepers Anne A Glumoff Tuomo T Kellokumpu Sakari S

The Journal of biological chemistry 20190808 39

β-1,4-Galactosyltransferase 1 (B4GALT1) and ST6 β-galactoside α-2,6-sialyltransferase 1 (ST6GAL1) catalyze the successive addition of terminal β-1,4-linked galactose and α-2,6-linked sialic acid to <i>N-</i>glycans. Their exclusive interaction in the Golgi compartment is a prerequisite for their full catalytic activity, whereas a lack of this interaction is associated with cancers and hypoxia. To date, no structural information exists that shows how glycosyltransferases functionally assemble wit ...[more]

PMID: 31395657

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Project description:Epigenetic gene silencing is of central importance to maintain genome integrity and is mediated by an elaborate interplay between DNA methylation, histone posttranslational modifications and chromatin remodeling complexes. DNA methylation and repressive histone marks usually correlate with transcriptionally silent heterochromatin, however there are exceptions to this interdependence. In Arabidopsis, mutation of MORPHEUS MOLECULE 1 (MOM1) causes transcriptional derepression of heterochromatin independently of changes in DNA methylation. More recently, two Arabidopsis homologs of mouse Microrchidia (MORC) have also been implicated in gene silencing and heterochromatin condensation without altering genome-wide DNA methylation patterns. In this study, we show that AtMORC6 physically interacts with AtMORC1 and with its close homologue AtMORC2 in two mutually exclusive protein complexes. RNA-seq analysis of high-order mutants indicates that AtMORC1 and AtMORC2 act redundantly to repress a common set of loci. We also examined the genetic interactions between AtMORC6 and MOM1 pathways. Although AtMORC6 and MOM1 control the silencing of a very similar set of genomic loci, we observed synergistic transcriptional regulation in the mom1/atmorc6 double mutant, suggesting that these epigenetic regulators act mainly by independent silencing mechanisms. RNA-seq libraries were prepared for two suites of mutants to allow direct comparisons between mutants within each set. The two sets consisted of the following samples: Set_1) A wildtype (Col) control, the morc1 mutant, the morc2 mutant, the morc1 morc2 double mutant, the morc6 mutant, and the morc1 morc2 morc6 triple mutant ; Set_2) A wildtpe (Col) control, the morc6 mutant, the mom1 mutant, and the mom1 morc6 double mutant. For each sample, two biological replicates were performed (denoted "bio_replicate_1" or "bio_replicate_2"). Whole genome bisulifte libraries were sequenced from material grown in parallel.

Dataset Information

Assembly of B4GALT1/ST6GAL1 heteromers in the Golgi membranes involves lateral interactions via highly charged surface domains.

Publications

Assembly of B4GALT1/ST6GAL1 heteromers in the Golgi membranes involves lateral interactions via highly charged surface domains.

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