Analysis of transport and targeting of syndecan-1: effect of cytoplasmic tail deletions.

Abstract

Madin-Darby canine kidney (MDCK) cells and Chinese hamster ovary (CHO) cells were transfected with wild-type and cytoplasmic deletion mutants of mouse syndecan-1 to study the requirements for transport and polarized expression of this proteoglycan. Expression in MDCK cells revealed that wild-type syndecan-1 is directed to the basolateral surface via a brefeldin A-insensitive route. A deletion of the last 12 amino acids of the syndecan-1 cytoplasmic tail (CT22) was sufficient to result in the appearance of mutant proteoglycans at both the basolateral and apical cell surfaces. Treatment with brefeldin A was able to prevent apical transport of the mutants. We thus propose that the C-terminal part of the cytoplasmic tail is required for steady-state basolateral distribution of syndecan-1. In CHO cells a deletion of the last 25 or 33 amino acids of the 34-residue cytoplasmic domain (CT9 and CT1, respectively) resulted in partial retention of the mutants in the endoplasmic reticulum (ER). A deletion mutant lacking the last 12 amino acids (CT22) was not retained. Interestingly, the unglycosylated core proteins of the CT9 and CT1 mutants showed a significantly lower apparent molecular weight when analyzed by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis than wild-type syndecan-1. However, when CHO transfectants expressing the CT1 mutant were incubated with brefeldin A, causing fusion of the ER and Golgi, CT1 ran with an almost equally high apparent molecular weight as the wild-type molecule. This would suggest that syndecan-1 undergoes extensive posttranslational modifications or forms an SDS-resistant dimer/complex after transit from the ER.

Syndecans: multifunctional cell-surface co-receptors

“…Surprisingly, although syndecan-1 molecules that lack the cytoplasmic domain do not associate with actin filaments, they are not extractable by Triton X-100 after antibody-mediated clustering. These results, together with a previous report on low-pH-induced syndecan aggregation [137], demonstrate that detergent-insolubility of syndecan-1 results from a process that is not related to cytoskeleton association.…”

“…Incubation of cells in low-pH medium (pH 5) results in the conversion of syndecan-1 into a form that is not extractable by non-ionic detergent. Detergent-insolubility of the syndecan-1 molecules in these cells is not dependent on the cytoplasmic domain, but, instead, on the heparan sulphate chains [137]. Thus detergent-insolubility of syndecan-1 appears to result from an extracellular interaction, and not from an intracellular (i.e.…”

“…Expression of syndecan-1 in transfected Madin-Darby canine kidney cells results in the targeting of the proteoglycan to the basolateral surface. Deletion of the C-terminal 12 amino acids of the cytoplasmic domain results in the appearance of syndecan-1 in both the basolateral and apical compartments of the plasma membrane [72].…”

Evaluation of the Association with Cervical Cancer of Polymorphisms in Syndecan-1, a Heparan Sulfate Proteoglycan Involved with Viral Cell Entry

“…Although little is currently known about the expression of SDC-1 in different histologic compartments of the cervix, there is some evidence which indicates that different heparan sulfates may be found on different cell surfaces (e.g., SDC in basolateral compartments; refs. [10][11][12]. Despite the lack of direct data on differential expression of SDC-1 in different types of cervical cells, it is possible that there might be specific localization that ties to disease risk.…”

The Role of Glycans in Apical Sorting of Proteins in Polarized Epithelial Cells

“…Basolateral localization of syndecan-1 in MDCK cells requires a signal positioned in the 12 most distal amino acids of its cytoplasmic tail (129), while basolateral secretion of the major basement membrane heparan sulfate (HS) PG occurs by a pH-dependent sorting mechanism (130). The fact that different types of proteoglycans are secreted apically and basolaterally indicates that these molecules are actively sorted.…”