UV irradiation-induced production of monoglycosylated biglycan through downregulation of xylosyltransferase 1 in cultured human dermal fibroblasts.
BACKGROUND
Biglycan (BGN) is a proteoglycan composed of a 42-kDa core protein and two glycosaminoglycan (GAG) chains, and known to be involved in structural, space-filling functions and many physiological regulations in the skin.


OBJECTIVE
To investigate ultraviolet (UV) irradiation-induced changes of BGN protein and its GAG chain synthesis in cultured human dermal fibroblasts.


METHODS
UV irradiation-induced or xylosyltransferase (XYLT) 1 siRNA-mediated smaller-sized protein bands detected by Western blot using BGN antibodies were identified as monoglycosylated forms of BGN, using BGN siRNA-mediated knockdown and chondroitinase ABC (ChABC). Differential activity of XYLT1 and 2 on BGN core protein was investigated by size shift of S42A- and S47A-BGN mutants to core protein size caused by XYLT1 siRNA transfection or UV irradiation.


RESULTS
After UV irradiation, intact form of BGN protein (I-BGN) and core protein form were reduced in cultured fibroblasts, but other smaller-sized bands were observed to be increased. These smaller-sized ones were reduced by transfection of BGN siRNA, and shifted to the core protein size by treatment with ChABC, suggesting that they are defectively-glycosylated forms of BGN (D-BGN) protein. UV irradiation also decreased mRNA expression levels of XYLT1 and 2, which are responsible for initiation of GAG chain synthesis. UV-mediated reduction of XYLT1 expression was much stronger than that of XYLT2. Furthermore, siRNA-mediated down-regulation of XYLT1 resulted in the increase of D-BGN and the decrease of I-BGN, while down-regulation of XYLT2 resulted in no change of D-BGN and I-BGN, suggesting that the XYLT1 may react with both GAG-attaching serine sites of BGN; however, XYLT2 may prefer to react one of them. Another dermatan sulfate (DS) proteoglycan, decorin, showed no or a little change of its molecular weight by UV irradiation or XYLT1 siRNA transfection, suggesting that DS synthesis may not be a critical factor in formation of D-BGN. Co-transfection with XYLT1, 2 siRNAs and wild-type or mutant forms of BGN overexpression vectors revealed that S42A-BGN showed size reduction to core protein size by XYLT1 downregulation, but S47A-BGN did not, suggesting that XYLT2 can react only with S42 on BGN core protein. With UV irradiation, both S42A-BGN and S47A-BGN showed size reduction, which is probably because UV-caused downregulation of both XYLTs and overexpression condition resulted in incomplete glycosylation and secretion.


CONCLUSIONS
UV irradiation-induced increase of BGN monoglycosylated forms in cultured human dermal fibroblasts is resulted from dominance of XYLT2 activity, which acts only at S42 on BGN core protein, caused by UV-mediated stronger reduction of XYLT1.
Department of Dermatology, Seoul National University College of Medicine, Seoul, Republic of Korea; Department of Dermatology, Yanbian University Hospital, Yanji, Jilin, China; Laboratory of Cutaneous Aging Research, Biomedical Research Institute, Seoul National University Hospital, Seoul, Republic of Korea; Institute of Human-Environment Interface Biology, Medical Research Center, Seoul National University, Seoul, Republic of Korea. Electronic address: 15526770921@163.com.
Cheng Long
CL
Jin
Department of Dermatology, Seoul National University College of Medicine, Seoul, Republic of Korea; Laboratory of Cutaneous Aging Research, Biomedical Research Institute, Seoul National University Hospital, Seoul, Republic of Korea; Institute of Human-Environment Interface Biology, Medical Research Center, Seoul National University, Seoul, Republic of Korea.
Jang-Hee
JH
Oh
Department of Dermatology, Seoul National University College of Medicine, Seoul, Republic of Korea; Laboratory of Cutaneous Aging Research, Biomedical Research Institute, Seoul National University Hospital, Seoul, Republic of Korea; Institute of Human-Environment Interface Biology, Medical Research Center, Seoul National University, Seoul, Republic of Korea.
Mira
M
Han
Department of Dermatology, Seoul National University College of Medicine, Seoul, Republic of Korea; Laboratory of Cutaneous Aging Research, Biomedical Research Institute, Seoul National University Hospital, Seoul, Republic of Korea; Institute of Human-Environment Interface Biology, Medical Research Center, Seoul National University, Seoul, Republic of Korea.
Min Kyeong
MK
Shin
Department of Dermatology, Seoul National University College of Medicine, Seoul, Republic of Korea; Laboratory of Cutaneous Aging Research, Biomedical Research Institute, Seoul National University Hospital, Seoul, Republic of Korea; Institute of Human-Environment Interface Biology, Medical Research Center, Seoul National University, Seoul, Republic of Korea.
Cheng
C
Yao
Department of Dermatology, Seoul National University College of Medicine, Seoul, Republic of Korea; Laboratory of Cutaneous Aging Research, Biomedical Research Institute, Seoul National University Hospital, Seoul, Republic of Korea; Institute of Human-Environment Interface Biology, Medical Research Center, Seoul National University, Seoul, Republic of Korea.
Chi-Hyun
CH
Park
Department of Dermatology, Yanbian University Hospital, Yanji, Jilin, China. Electronic address: jinzh_621@163.com.
Zhe Hu
ZH
Jin
Department of Dermatology, Seoul National University College of Medicine, Seoul, Republic of Korea; Laboratory of Cutaneous Aging Research, Biomedical Research Institute, Seoul National University Hospital, Seoul, Republic of Korea; Institute of Human-Environment Interface Biology, Medical Research Center, Seoul National University, Seoul, Republic of Korea. Electronic address: jhchung@snu.ac.kr.
Jin Ho
JH
Chung
D058578:Biglycan / Q000096:biosynthesis* / Q000235:genetics*; D002478:Cells, Cultured; D058575:Decorin / Q000378:metabolism; D015536:Down-Regulation / Q000528:radiation effects; D005347:Fibroblasts / Q000378:metabolism / Q000528:radiation effects; D020868:Gene Silencing; D006025:Glycosaminoglycans / Q000096:biosynthesis* / Q000528:radiation effects; D006031:Glycosylation / Q000528:radiation effects; D006801:Humans; D008970:Molecular Weight; D010430:Pentosyltransferases / Q000235:genetics / Q000378:metabolism* / Q000528:radiation effects; D014176:Protein Biosynthesis / Q000528:radiation effects; D012333:RNA, Messenger / Q000378:metabolism; D012879:Skin Physiological Phenomena / Q000528:radiation effects; D014466:Ultraviolet Rays; D014994:Xylose / Q000378:metabolism; D000097608:UDP Xylose-Protein Xylosyltransferase
D058578:Biglycan; D058575:Decorin; D006025:Glycosaminoglycans; D010430:Pentosyltransferases; D012333:RNA, Messenger; D014994:Xylose; D000097608:UDP Xylose-Protein Xylosyltransferase
Biglycan; Fibroblast; UV; XYLT1; XYLT2