Limits...
Mutants in trs120 disrupt traffic from the early endosome to the late Golgi.

Cai H, Zhang Y, Pypaert M, Walker L, Ferro-Novick S - J. Cell Biol. (2005)

Bottom Line: Transport protein particle (TRAPP), a large complex that mediates membrane traffic, is found in two forms (TRAPPI and -II).Surprisingly, we report that mutations in trs120 do not block general secretion.Furthermore, we demonstrate that Trs120p largely colocalizes with the late Golgi marker Sec7p.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, CT 06519, USA.

ABSTRACT
Transport protein particle (TRAPP), a large complex that mediates membrane traffic, is found in two forms (TRAPPI and -II). Both complexes share seven subunits, whereas three subunits (Trs130p, -120p, and -65p) are specific to TRAPPII. Previous studies have shown that mutations in the TRAPPII-specific gene trs130 block traffic through or from the Golgi. Surprisingly, we report that mutations in trs120 do not block general secretion. Instead, trs120 mutants accumulate aberrant membrane structures that resemble Berkeley bodies and disrupt the traffic of proteins that recycle through the early endosome. Mutants defective in recycling also display a defect in the localization of coat protein I (COPI) subunits, implying that Trs120p may participate in a COPI-dependent trafficking step on the early endosomal pathway. Furthermore, we demonstrate that Trs120p largely colocalizes with the late Golgi marker Sec7p. Our findings imply that Trs120p is required for vesicle traffic from the early endosome to the late Golgi.

Show MeSH

Related in: MedlinePlus

Mutants in trs120 do not block general secretion. Cells were grown to early log phase at 25°C, preincubated to 37°C for 20 min, radiolabeled for 15 min, and then chased for 30 min. The growth medium was separated from the cells by centrifugation, processed for TCA precipitation, and analyzed by SDS-PAGE on a 10% gel as described previously (Kim et al., 2001). The asterisks mark proteins secreted into the medium in wild-type (WT) cells.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2171297&req=5

fig3: Mutants in trs120 do not block general secretion. Cells were grown to early log phase at 25°C, preincubated to 37°C for 20 min, radiolabeled for 15 min, and then chased for 30 min. The growth medium was separated from the cells by centrifugation, processed for TCA precipitation, and analyzed by SDS-PAGE on a 10% gel as described previously (Kim et al., 2001). The asterisks mark proteins secreted into the medium in wild-type (WT) cells.

Mentions: The finding that trs120-2 and -8 mutants do not block the trafficking of CPY and only partially block invertase secretion prompted us to determine whether there is a general block in secretion in these mutants. To assay for a general secretion defect, we shifted wild-type and mutant cells to 37°C for 20 min, pulse labeled them for 15 min, and chased them for 30 min. Cells were pelleted, and the proteins secreted into the medium were precipitated with TCA and resolved on an SDS–polyacrylamide gel (Fig. 3). Surprisingly, although the sec18, trs130ts2, and trs130-6 mutants blocked secretion (Fig. 3, compare lane 1 with lanes 2, 5, and 6), no obvious defect was observed in the trs120-2 and -8 mutants (lanes 3 and 4). The block in secretion was complete in sec18 but not in trs130 mutants (see Fig. S1, available at http://www.jcb.org/cgi/content/full/jcb.200505145/DC1, for darker exposure of the autorad). Therefore, although mutations in the trs130 gene lead to a defect in the trafficking of proteins within the Golgi, the loss of trs120 function does not result in a significant block in secretion.


Mutants in trs120 disrupt traffic from the early endosome to the late Golgi.

Cai H, Zhang Y, Pypaert M, Walker L, Ferro-Novick S - J. Cell Biol. (2005)

Mutants in trs120 do not block general secretion. Cells were grown to early log phase at 25°C, preincubated to 37°C for 20 min, radiolabeled for 15 min, and then chased for 30 min. The growth medium was separated from the cells by centrifugation, processed for TCA precipitation, and analyzed by SDS-PAGE on a 10% gel as described previously (Kim et al., 2001). The asterisks mark proteins secreted into the medium in wild-type (WT) cells.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171297&req=5

fig3: Mutants in trs120 do not block general secretion. Cells were grown to early log phase at 25°C, preincubated to 37°C for 20 min, radiolabeled for 15 min, and then chased for 30 min. The growth medium was separated from the cells by centrifugation, processed for TCA precipitation, and analyzed by SDS-PAGE on a 10% gel as described previously (Kim et al., 2001). The asterisks mark proteins secreted into the medium in wild-type (WT) cells.
Mentions: The finding that trs120-2 and -8 mutants do not block the trafficking of CPY and only partially block invertase secretion prompted us to determine whether there is a general block in secretion in these mutants. To assay for a general secretion defect, we shifted wild-type and mutant cells to 37°C for 20 min, pulse labeled them for 15 min, and chased them for 30 min. Cells were pelleted, and the proteins secreted into the medium were precipitated with TCA and resolved on an SDS–polyacrylamide gel (Fig. 3). Surprisingly, although the sec18, trs130ts2, and trs130-6 mutants blocked secretion (Fig. 3, compare lane 1 with lanes 2, 5, and 6), no obvious defect was observed in the trs120-2 and -8 mutants (lanes 3 and 4). The block in secretion was complete in sec18 but not in trs130 mutants (see Fig. S1, available at http://www.jcb.org/cgi/content/full/jcb.200505145/DC1, for darker exposure of the autorad). Therefore, although mutations in the trs130 gene lead to a defect in the trafficking of proteins within the Golgi, the loss of trs120 function does not result in a significant block in secretion.

Bottom Line: Transport protein particle (TRAPP), a large complex that mediates membrane traffic, is found in two forms (TRAPPI and -II).Surprisingly, we report that mutations in trs120 do not block general secretion.Furthermore, we demonstrate that Trs120p largely colocalizes with the late Golgi marker Sec7p.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, CT 06519, USA.

ABSTRACT
Transport protein particle (TRAPP), a large complex that mediates membrane traffic, is found in two forms (TRAPPI and -II). Both complexes share seven subunits, whereas three subunits (Trs130p, -120p, and -65p) are specific to TRAPPII. Previous studies have shown that mutations in the TRAPPII-specific gene trs130 block traffic through or from the Golgi. Surprisingly, we report that mutations in trs120 do not block general secretion. Instead, trs120 mutants accumulate aberrant membrane structures that resemble Berkeley bodies and disrupt the traffic of proteins that recycle through the early endosome. Mutants defective in recycling also display a defect in the localization of coat protein I (COPI) subunits, implying that Trs120p may participate in a COPI-dependent trafficking step on the early endosomal pathway. Furthermore, we demonstrate that Trs120p largely colocalizes with the late Golgi marker Sec7p. Our findings imply that Trs120p is required for vesicle traffic from the early endosome to the late Golgi.

Show MeSH
Related in: MedlinePlus