The sulfation of tyrosine residues is an important posttranslational modification involved in the regulation of protein function. We examined the activity of worm tyrosylprotein sulfotransferase (TPST-1) on a typical cuticle collagen, ROL-6, in C. elegans. We verified that TPST-1 sulfates three tyrosine residues of ROL-6 in vitro. We found that these tyrosine residues are important for the secretion of ROL-6::GFP. Mutant ROL-6::GFP proteins that contain more than two substitutions of the target tyrosine residues are severely deficient in cuticle localization. Consistently, knock down of tpst-1 blocked the cuticle localization of ROL-6::GFP. Therefore, the sulfation of ROL-6 by TPST-1 is critical for the proper localization of ROL-6. We also confirmed that worm TPST-1 is localized to the trans-Golgi network (TGN). Our results indicate that TPST-1 regulates cuticle organization by promoting the transport of ROL-6 from the TGN to the cuticle.