A mutation in signal peptide of rat resistin gene inhibits differentiation of 3T3-L1 preadipocytes.
OBJECTIVE
To detect the resistin expression of white adipose tissue in diet-induced obese (DIO) versus diet-resistant (DR) rats, and to investigate the relationship of mutated resistin and 3T3-L1 preadipocytes differentiation.


METHODS
RT-PCR and Western Blot were used to detect gene /protein expression. 3T3-L1 cells were cultured, transfected, and induced to differentiation using 0.5 mmol/L 3-isobutyl-1-methylxanthine (MIX), 1 mg/L insulin, and 1 micromol/L dexamethasone. Oil red O staining was applied to detect the degree of preadipocytes differentiation.


RESULTS
Expression of resistin mRNA was upregulated in DIO rats and downregulated in DR rats. However, the expression levels varied greatly within the groups. Sequencing of the resistin genes from DIO and DR rats revealed a Leu9Val (C25G) missense mutation within the signal peptide in one DR rat. The mutant resistin inhibited preadipocyte differentiation. Local experiments and Western blotting with tagged resistin fusion proteins identified both mutant and wild type proteins in the cytoplasm and secreted into the culture medium. Computer predictions using the Proscan and Subloc programs revealed four putative phosphorylation sites and a possible leucine zipper motif within the rat resistin protein.


CONCLUSIONS
Resistin-increased differentiation may be inhibited by the mutation-containing precursor protein, or by the mutant non-secretory resistin isoform.
Guo
Xi-rong
XR
Department of Pediatrics, Center of Human Functional Genomics, Nanjing Medical University, Nanjing 210029, China.
Gong
Hai-xia
HX
Gao
Yan-qin
YQ
Fei
Li
L
Ni
Yu-hui
YH
Chen
Rong-hua
RH
D041721:3T3-L1 Cells; D017667:Adipocytes / Q000166:cytology* / Q000187:drug effects; D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D004041:Dietary Fats; D015870:Gene Expression / Q000187:drug effects; D005822:Genetic Vectors; D006729:Hormones, Ectopic / Q000235:genetics* / Q000494:pharmacology; D008297:Male; D051379:Mice; D020125:Mutation, Missense; D009765:Obesity / Q000235:genetics / Q000378:metabolism; D021382:Protein Sorting Signals / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D052243:Resistin; D014162:Transfection
D004041:Dietary Fats; D006729:Hormones, Ectopic; D021382:Protein Sorting Signals; D012333:RNA, Messenger; D052243:Resistin; C499623:Retn protein, mouse; C499622:Retn protein, rat