Fidgetin-like 1 gene inhibited by basic fibroblast growth factor regulates the proliferation and differentiation of osteoblasts.
The FIGNL1 gene was proven to be a new subfamily member of ATPases associated with diverse cellular activities (AAA proteins). In this in vitro study, the AAA proteins inhibited osteoblast proliferation and stimulated osteoblast differentiation. We showed that FIGNL1 may play some regulatory role in osteoblastogenesis.


BACKGROUND
The fidgetin-like 1 (FIGNL1) gene encodes a new subfamily member of ATPases associated with diverse cellular activities (AAA proteins). Although the FIGNL1 protein localizes to both the nucleus and cytoplasm, the function of FIGNL1 remains unknown. In a previous study, we identified several genes that mediate the anabolic effects of basic fibroblast growth factor (bFGF) on bone by using microarray data. FIGNL1 was one of the genes that downregulated >2-fold in MC3T3-E1 cells after treatment with bFGF. Therefore, this study was aimed to identify and confirm the function of FIGNL1 on osteoblastogenesis.


METHODS
We examined the effect of the FIGNL1 gene on proliferation, differentiation, and apoptosis in mouse osteoblast cells (MC3T3-E1 and mouse primary calvarial cells) using flow cytometry, RT-PCR, cell proliferation assay, and cell death assay. MC3T3-E1 cells and mouse calvarial cells were transfected with small interfering RNA (siRNA) directed against the FIGNL1 or nontargeting control siRNA and examined by cell proliferation and cell death assays. Also, FIGNL1 was fused to enhance green fluorescent protein (EGFP), and the EGFP-fused protein was transiently expressed in MC3T3-E1 cells.


RESULTS
Reduced expression of FIGNL1 by bFGF and TGF-beta1 treatment was verified by RT-PCR analysis. Overexpression of FIGNL1 reduced the proliferation of MC3T3-E1 and calvarial cells, more than the mock transfected control cells did. In contrast, siFIGNL1 transfection significantly increased the proliferation of osteoblasts, whereas overexpression of FIGNL1 did not seem to alter apoptosis in osteoblasts. Meanwhile, overexpression of FIGNL1 enhanced the mRNA expression of alkaline phosphatase (ALP) and osteocalcin (OCN) in osteoblasts. In contrast, siFIGNL1 decreased the expression of ALP and OCN. A pEGFP-FIGNL1 transfected into MCT3-E1 cells had an initially ubiquitous distribution and rapidly translocated to the nucleus 1 h after bFGF treatment.


CONCLUSIONS
From these results, we proposed that FIGNL1, a subfamily member of the AAA family of proteins, might play some regulatory role in osteoblast proliferation and differentiation. Further analyses of FIGNL1 will be needed to better delineate the mechanisms contributing to the inhibition of proliferation and stimulation of osteoblast differentiation.
Park
Su Jin
SJ
Brain Korea 21 Project for Medical Science, College of Medicine, Yonsei University, Seoul, Republic of Korea.
Kim
Su Jin
SJ
Rhee
Yumie
Y
Byun
Ji Hyun
JH
Kim
Seong Hwan
SH
Kim
Myoung Hee
MH
Lee
Eun Jig
EJ
Lim
Sung-Kil
SK
D016475:3T3 Cells; D000074183:ATPases Associated with Diverse Cellular Activities; D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D000469:Alkaline Phosphatase / Q000235:genetics; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics / Q000502:physiology; D002452:Cell Count; D002454:Cell Differentiation / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D024042:Collagen Type I / Q000235:genetics; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008813:Mice, Inbred ICR; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D015675:Osteocalcin / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D012886:Skull / Q000166:cytology; D014162:Transfection; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology
D024042:Collagen Type I; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D053773:Transforming Growth Factor beta1; D016222:Fibroblast Growth Factor 2; D015675:Osteocalcin; D049452:Green Fluorescent Proteins; D000469:Alkaline Phosphatase; D000251:Adenosine Triphosphatases; D000074183:ATPases Associated with Diverse Cellular Activities; C522287:FIGNL1 protein, mouse; C498542:Fign protein, mouse