Brickner J H, Fuller R S
Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor 48109-0606, USA.
J Cell Biol. 1997 Oct 6;139(1):23-36. doi: 10.1083/jcb.139.1.23.
Localization of yeast Kex2 protease to the TGN requires a signal (TLS1) in its cytosolic tail (C-tail). Mutation of TLS1 results in rapid transit of Kex2p to the vacuole. Isolation of suppressors of the Tyr713Ala mutation in TLS1 previously identified three SOI genes. SOI1, cloned by complementation of a sporulation defect, encodes a novel, hydrophilic 3,144-residue protein with homologues in Caenorhabditis elegans, Drosophila melanogaster, and humans. Epitope-tagged Soi1p existed in a detergent-insensitive, sedimentable form. Deletion of SOI1 impaired TGN localization of wild-type Kex2p and a fusion protein containing the C-tail of Ste13p, and also caused missorting of carboxypeptidase Y and accelerated vacuolar degradation of the Vps10p sorting receptor. Deletion of SOI1 improved retention of Tyr713Ala Kex2p in the pro-alpha-factor processing compartment but, unlike the original soi1 alleles, did not increase the half-life of Tyr713Ala Kex2p. These results suggested that Soi1p functions at two steps in the cycling of Kex2p and other proteins between the TGN and prevacuolar compartment (PVC). This hypothesis was confirmed in several ways. Soi1p was shown to be required for optimal function of TLS1. Suppression of the Tyr713Ala mutation by mutation of SOI1 was shown to be caused by activation of a second signal (TLS2) in the Kex2p C-tail. TLS2 delayed exit of Kex2p from the TGN, whereas TLS1 did not affect this step. We propose that Soi1p promotes cycling of TGN membrane proteins between the TGN and PVC by antagonizing a TGN retention signal (TLS2) and facilitating the function of a retrieval signal (TLS1) that acts at the PVC.
酵母Kex2蛋白酶定位于反式高尔基体网络（TGN）需要其胞质尾（C尾）中的一个信号（TLS1）。TLS1的突变导致Kex2p快速转运至液泡。先前对TLS1中Tyr713Ala突变的抑制子进行分离鉴定出了三个SOI基因。通过互补孢子形成缺陷而克隆的SOI1编码一种新型的亲水性3144个残基的蛋白质，该蛋白质在秀丽隐杆线虫、果蝇和人类中存在同源物。表位标记的Soi1p以一种对去污剂不敏感、可沉降的形式存在。SOI1的缺失损害了野生型Kex2p和包含Ste13p C尾的融合蛋白的TGN定位，还导致羧肽酶Y分选错误并加速Vps10p分选受体的液泡降解。SOI1的缺失改善了Tyr713Ala Kex2p在前体α因子加工区室中的保留，但与原始的soi1等位基因不同，并未增加Tyr713Ala Kex2p的半衰期。这些结果表明，Soi1p在Kex2p和其他蛋白质在TGN和前液泡区室（PVC）之间循环的两个步骤中发挥作用。这一假设通过多种方式得到了证实。结果表明，Soi1p是TLS1最佳功能所必需的。SOI1突变对Tyr713Ala突变的抑制作用被证明是由Kex2p C尾中的第二个信号（TLS2）激活所致。TLS2延迟了Kex2p从TGN的输出，而TLS1不影响这一步骤。我们提出，Soi1p通过拮抗TGN保留信号（TLS2）并促进在PVC起作用的回收信号（TLS1）的功能，来促进TGN膜蛋白在TGN和PVC之间的循环。
