DIFFERENT SENSITIVITY TO WORTMANNIN OF 2 VACUOLAR SORTING SIGNALS INDICATES THE PRESENCE OF DISTINCT SORTING MACHINERIES IN TOBACCO CELLS

被引:215
作者
MATSUOKA, K [1 ]
BASSHAM, DC [1 ]
RAIKHEL, NV [1 ]
NAKAMURA, K [1 ]
机构
[1] MICHIGAN STATE UNIV,DOE,PLANT RES LAB,E LANSING,MI 48824
关键词
D O I
10.1083/jcb.130.6.1307
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
Vacuolar matrix proteins in plant cells are sorted from the secretory pathway to the vacuoles at the Golgi apparatus. Previously, we reported that the NH2-terminal propeptide (NTPP) of the sporamin precursor and the COOH-terminal propeptide (CTPP) of the barley lectin precursor contain information for vacuolar sorting. To analyze whether these propeptides are interchangeable, we expressed constructs consisting of wild-type or mutated NTPP with the mature part of barley lectin and sporamin with CTPP and mutated NTPP in tobacco BY-2 cells. The vacuolar localization of these constructs indicated that the signals were interchangeable. We next analyzed the effect of wortmannin, a specific inhibitor of mammalian phosphatidylinositol (PI) 3-kinase on vacuolar delivery by NTPP and CTPP in tobacco cells. Pulse-chase analysis indicated that 33 mu M wortmannin caused almost complete inhibition of CTPP-mediated transport to the vacuoles, while NTPP-mediated transport displayed almost no sensitivity to wortmannin at this concentration. This indicates that there are at least two different mechanisms for vacuolar sorting in tobacco cells, and the CTPP-mediated pathway is sensitive to wortmannin. We compared the dose dependencies of wortmannin on the inhibition of CTPP-mediated vacuolar delivery of proteins and on the inhibition of the synthesis of phospholipids in tobacco cells. Wortmannin inhibited PI 3- and PI 4-kinase activities and phospholipid synthesis. Missorting caused by wortmannin displays a dose dependency that is similar to the dose dependency for the inhibition of synthesis of PI 4-phosphate and major phospholipids. This is different, however, than the inhibition of synthesis of PI 3-phosphate. Thus, the synthesis of phospholipids could be involved in CTPP-mediated vacuolar transport.
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页码:1307 / 1318
页数:12
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