The nucleotide sequence of the short and long transcripts of beta1,4- N -acetylgalactosaminyltransferase have been submitted to the DDBJ, EMBL, GenBank(R) and GSDB Nucleotide Sequence Databases under accession nos AJ517770 and AJ517771 respectively. The human Sd(a) antigen is formed through the addition of an N -acetylgalactosamine residue via a beta1,4-linkage to a sub-terminal galactose residue substituted with an alpha2,3-linked sialic acid residue. We have taken advantage of the previously cloned mouse cDNA sequence of the UDP-GalNAc:Neu5Acalpha2-3Galbeta-R beta1,4- N -acetylgalactosaminyltransferase (Sd(a) beta1,4GalNAc transferase) to screen the human EST and genomic databases and to identify the corresponding human gene. The sequence spans over 35 kb of genomic DNA on chromosome 17 and comprises at least 12 exons. As judged by reverse transcription PCR, the human gene is expressed widely since it is detected in various amounts in almost all cell types studied. Northern blot analysis indicated that five Sd(a) beta1,4GalNAc transferase transcripts of 8.8, 6.1, 4.7, 3.8 and 1.65 kb were highly expressed in colon and to a lesser extent in kidney, stomach, ileum and rectum. The complete coding nucleotide sequence was amplified from Caco-2 cells. Interestingly, the alternative use of two first exons, named E1(S) and E1(L), leads to the production of two transcripts. These nucleotide sequences give rise potentially to two proteins of 506 and 566 amino acid residues, identical in their sequence with the exception of their cytoplasmic tail. The short form is highly similar (74% identity) to the mouse enzyme whereas the long form shows an unusual long cytoplasmic tail of 66 amino acid residues that is as yet not described for any other mammalian glycosyltransferase. Upon transient transfection in Cos-7 cells of the common catalytic domain, a soluble form of the protein was obtained, which catalysed the transfer of GalNAc residues to alpha2,3-sialylated acceptor substrates, to form the GalNAcbeta1-4[Neu5Acalpha2-3]Galbeta1-R trisaccharide common to both Sd(a) and Cad antigens.

We have examined the hypothesis that the highly selective recombination of an active mating type locus (MAT) with either HML␣ or HMRa is facilitated by the spatial positioning of relevant sequences within the budding yeast (Saccharomyces cerevisiae) nucleus. However, both position relative to the nuclear envelope (NE) and the subnuclear mobility of fluorescently tagged MAT, HML, or HMR loci are largely identical in haploid a and ␣ cells. Irrespective of mating type, the expressed MAT locus is highly mobile within the nuclear lumen, while silent loci move less and are found preferentially near the NE. The perinuclear positions of HMR and HML are strongly compromised in strains lacking the Silent information regulator, Sir4. However, HML␣, unlike HMRa and most telomeres, shows increased NE association in a strain lacking yeast Ku70 (yKu70). Intriguingly, we find that the yKu complex is associated with HML and HMR sequences in a mating-typespecific manner. Its abundance decreases at the HML␣ donor locus and increases transiently at MATa following DSB induction. Our data suggest that mating-type-specific binding of yKu to HML␣ creates a local chromatin structure competent for recombination, which cooperates with the recombination enhancer to direct donor choice for gene conversion of the MATa locus.Long-range interactions between two genomic loci in distinct nuclear and chromatin environments are thought to influence recombination and transcription and to contribute to gene control during cell type differentiation in multicellular organisms (1,17,48,50). In Saccharomyces cerevisiae, a-or ␣-cell type is determined by two homeobox-containing genes transcribed at the MAT locus. The ability of yeast cells to switch mating type requires transcriptionally silent copies of both a and ␣ information, which are generally found at the homologous mating type loci HML and HMR (19). Genes at the HM loci, like genes in subtelomeric regions, are prone to position-dependent transcriptional repression. This repression is mediated by the recruitment and spreading of a Silent information regulatory (Sir) complex of Sir2, Sir3, and Sir4 (45).For subtelomeric genes, Sir-mediated repression is facilitated by the clustering of telomeres near the nuclear envelope (NE) (18, 21, 51), which generates a high local concentration of Sir factors. Since Sir factors are limiting for repression (32), the juxtaposition of a reporter gene near such clusters favors repression (2). Telomere anchoring itself is mediated by two redundant pathways: one that requires the yeast Ku70/Ku80 (yKu70/80) complex, and a second that is mediated by Sir4 interaction with the membrane-associated Enhancer of silent chromatin 1 (Esc1) (21, 51). In contrast to the repression that is enhanced by NE association, the interaction of active genes with nuclear pore complexes can increase transcript levels for some inducible genes, possibly by providing a barrier between active and inactive chromatin domains (1,48).In this study, we have examined whether there is cell-typespecifi...