FBXL3

FBXL3 is a gene in humans and mice that encodes the F-box/LRR-repeat protein 3 (FBXL3).[5][6] FBXL3 is a member of the F-box protein family, which constitutes one of the four subunits in the SCF ubiquitin ligase complex.[7]

FBXL3
Available structures PDB Ortholog search: PDBe RCSB List of PDB id codes 4I6J
Identifiers
Aliases	FBXL3, FBL3, FBL3A, FBXL3A, F-box and leucine-rich repeat protein 3, F-box and leucine rich repeat protein 3, IDDSFAS
External IDs	OMIM: 605653 MGI: 1354702 HomoloGene: 8127 GeneCards: FBXL3
Gene location (Human) Chr. Chromosome 13 (human)[1] Band 13q22.3 Start 76,992,598 bp[1] End 77,027,195 bp[1]
Gene location (Mouse) Chr. Chromosome 14 (mouse)[2] Band 14|14 E2.3 Start 103,080,239 bp[2] End 103,099,566 bp[2]
Gene ontology Molecular function • GO:0001948 protein binding • ubiquitin-protein transferase activity • GO:1904264, GO:1904822, GO:0090622, GO:0090302 ubiquitin protein ligase activity Cellular component • cytoplasm • ubiquitin ligase complex • SCF ubiquitin ligase complex • cell nucleus • cytosol • nuclear body Biological process • SCF-dependent proteasomal ubiquitin-dependent protein catabolic process • entrainment of circadian clock by photoperiod • regulation of circadian rhythm • protein destabilization • protein ubiquitination • rhythmic process • proteasome-mediated ubiquitin-dependent protein catabolic process • protein polyubiquitination • post-translational modification Sources:Amigo / QuickGO
Orthologs
Species	Human	Mouse
Entrez	26224	50789
Ensembl	ENSG00000005812	ENSMUSG00000022124
UniProt	Q9UKT7	Q8C4V4
RefSeq (mRNA)	NM_012158	NM_015822 NM_001347600 NM_001347601 NM_001360341 NM_001360342
RefSeq (protein)	NP_036290	NP_001334529 NP_001334530 NP_056637 NP_001347270 NP_001347271
Location (UCSC)	Chr 13: 76.99 – 77.03 Mb	Chr 14: 103.08 – 103.1 Mb
PubMed search	[3]	[4]
Wikidata
View/Edit Human View/Edit Mouse

The FBXL3 protein participates in the negative feedback loop responsible for generating molecular circadian rhythms in mammals by binding to the CRY1 and CRY2 proteins to facilitate their polyubiquitination by the SCF complex and their subsequent degradation by the proteasome.[8][9][10]

Discovery

The Fbxl3 gene function was independently identified in 2007 by three groups lead by Michele Pagano, Joseph S. Takahashi, Dr. Patrick Nolan and Michael Hastings, respectively. Takahashi used forward genetics N-ethyl-N-nitrosourea (ENU) mutagenesis to screen for mice with varied circadian activity which led to the discovery of the Overtime (Ovtm) mutant of the Fbxl3 gene.[9] Nolan discovered the Fbxl3 mutation After hours (Afh) by a forward screen assessing wheel activity behavior of mutagenized mice.[10] The phenotypes identified in mice were mechanistically explained by Pagano who discovered that the FBXL3 protein is necessary for the reactivation of the CLOCK and BMAL1 protein heterodimer by inducing the degradation of CRY proteins.[8]

Overtime

Mice with the homozygous mutation of Ovtm, free run with an intrinsic period of 26 hours. Overtime is a loss of function mutation caused by a substitution of isoleucine to threonine in the region of FBXL3 that binds to CRY. In mice with this mutation, levels of the proteins PER1 and PER2 are decreased, while levels of CRY proteins do not differ from those of wild type mice. The stabilization of CRY protein levels leads to continued repression of Per1 and Per2 transcription and translation.[9]

After-hours

The After-hours mutation is a substitution of cysteine to serine at position 358. Similar to Overtime, the mutation occurs in the region where FBXL3 binds to CRY. Mice homozygous for the Afh mutation have a free running period of about 27 hours. The Afh mutation delays the rate of CRY protein degradation, therefore affecting the transcription of PER2 protein.[8][10]

Fbxl21

The closest homologue to Fbxl3 is Fbxl21 as it also binds to the CRY1 and CRY2 proteins. Predominantly localized to the cytosol, Fbxl21 has been proposed to antagonize the action of Fbxl3 through ubiquitination and stabilization of CRY proteins instead of leading it to degradation.[11] FBXL21 is expressed predominantly in the suprachiasmatic nucleus, which is the region in the brain that functions as the master pacemaker in mammals.[12]

Characteristics

The human FBXL3 gene is located on the long arm of chromosome 13 at position 22.3.[11][13] The protein is composed of 428 amino acids and has a mass of 48,707 Daltons.[14] The FBXL3 protein contains an F-box domain, characterized by a 40 amino acid motif that mediates protein-protein interactions, and several tandem leucine-rich repeats used for substrate recognition. It has eight post-translational modification sites involving ubiquitination and four sites involving phosphorylation. The FBXL3 protein is predominantly localized to the nucleus. It is one of four subunits of a ubiquitin ligase complex called SKP1-CUL1-F-box-protein, which includes the proteins CUL1, SKP1, and RBX1.[13][15]

Function

The FBXL3 protein plays a role in the negative feedback loop of the mammalian molecular circadian rhythm. The PER and CRY proteins inhibit the transcription factors CLOCK and BMAL1. The degradation of PER and CRY prevent the inhibition of the CLOCK and BMAL1 protein heterodimer. In the nucleus, the FBXL3 protein targets CRY1 and CRY2 for polyubiquitination, which triggers the degradation of the proteins by the proteasome.[8] FBXL3 binds to CRY2 by occupying its flavin adenine dinucleotide (FAD) cofactor pocket with a C-terminal tail and buries the PER-binding interface on the CRY2 protein.[16]

The FBXL3 protein is also involved in a related feedback loop that regulates the transcription of the Bmal1 gene. Bmal1 expression is regulated by the binding of REV-ERBα and RORα proteins to retinoic acid-related orphan receptor response elements (ROREs) in the Bmal1 promoter region. The binding of the REV-ERBα protein to the promoter represses expression, while RORα binding activates expression.[17] FBXL3 decreases the repression of Bmal1 transcription by inactivating the REV-ERBα and HDAC3 repressor complex.[18]

The FBXL3 protein has also been found to cooperatively degrade c-MYC when bound to CRY2. The c-MYC protein is a transcription factor important in regulating cell proliferation. The CRY2 protein can function as a co-factor for the FBXL3 ligase complex and interacts with phosphorylated c-MYC. This interaction promotes the ubiquitination and degradation of the c-MYC protein.[19]

Interactions

FBXL3 has been shown to interact with:

• SKP1A[13]
• CRY1 [17]
• CRY2 [19][20]
• REV-ERBα [18]
• HDAC3 [18]
• c-MYC [19]

References

1. GRCh38: Ensembl release 89: ENSG00000005812 - Ensembl, May 2017
2. GRCm38: Ensembl release 89: ENSMUSG00000022124 - Ensembl, May 2017
3. "Human PubMed Reference:". National Center for Biotechnology Information, U.S. National Library of Medicine.
4. "Mouse PubMed Reference:". National Center for Biotechnology Information, U.S. National Library of Medicine.
5. GRCh38: Ensembl release 89: ENSG00000005812 - Ensembl, May 2017
6. GRCm38: Ensembl release 89: ENSMUSG00000022124 - Ensembl, May 2017
7. "Human PubMed Reference:".
8. Busino L, Bassermann F, Maiolica A, Lee C, Nolan PM, Godinho SI, Draetta GF, Pagano M (May 2007). "SCFFbxl3 controls the oscillation of the circadian clock by directing the degradation of cryptochrome proteins". Science. 316 (5826): 900–4. doi:10.1126/science.1141194. PMID 17463251.
9. Siepka SM, Yoo SH, Park J, Lee C, Takahashi JS (2007). "Genetics and neurobiology of circadian clocks in mammals". Cold Spring Harbor Symposia on Quantitative Biology. 72: 251–259. doi:10.1101/sqb.2007.72.052. PMC 3749845. PMID 18419282.
10. Godinho SI, Maywood ES, Shaw L, Tucci V, Barnard AR, Busino L, Pagano M, Kendall R, Quwailid MM, Romero MR, O'neill J, Chesham JE, Brooker D, Lalanne Z, Hastings MH, Nolan PM (May 2007). "The after-hours mutant reveals a role for Fbxl3 in determining mammalian circadian period". Science. 316 (5826): 897–900. doi:10.1126/science.1141138. PMID 17463252.
11. Hirano A, Yumimoto K, Tsunematsu R, Matsumoto M, Oyama M, Kozuka-Hata H, Nakagawa T, Lanjakornsiripan D, Nakayama KI, Fukada Y (February 2013). "FBXL21 regulates oscillation of the circadian clock through ubiquitination and stabilization of cryptochromes". Cell. 152 (5): 1106–18. doi:10.1016/j.cell.2013.01.054. PMID 23452856.
12. Dardente H, Mendoza J, Fustin JM, Challet E, Hazlerigg DG (2008). "Implication of the F-Box Protein FBXL21 in circadian pacemaker function in mammals". PLOS ONE. 3 (10): e3530. doi:10.1371/journal.pone.0003530. PMC 2568807. PMID 18953409.
13. Cenciarelli C, Chiaur DS, Guardavaccaro D, Parks W, Vidal M, Pagano M (October 1999). "Identification of a family of human F-box proteins". Current Biology. 9 (20): 1177–9. doi:10.1016/S0960-9822(00)80020-2. PMID 10531035.
14. Sato K, Yoshida K (November 2010). "Augmentation of the ubiquitin-mediated proteolytic system by F-box and additional motif-containing proteins (Review)". International Journal of Oncology. 37 (5): 1071–6. doi:10.3892/ijo_00000758. PMID 20878054.
15. "FBXL3 F-box and leucine rich repeat protein 3 [ Homo sapiens (human) ]". Entrez Gene. Retrieved 27 April 2017.
16. Xing W, Busino L, Hinds TR, Marionni ST, Saifee NH, Bush MF, Pagano M, Zheng N (April 2013). "SCF(FBXL3) ubiquitin ligase targets cryptochromes at their cofactor pocket". Nature. 496 (7443): 64–8. doi:10.1038/nature11964. PMC 3618506. PMID 23503662.
17. Ko CH, Takahashi JS (October 2006). "Molecular components of the mammalian circadian clock". Human Molecular Genetics. 15 Spec No 2 (Review Issue 2): R271-7. doi:10.1093/hmg/ddl207. PMID 16987893.
18. Shi G, Xing L, Liu Z, Qu Z, Wu X, Dong Z, Wang X, Gao X, Huang M, Yan J, Yang L, Liu Y, Ptácek LJ, Xu Y (March 2013). "Dual roles of FBXL3 in the mammalian circadian feedback loops are important for period determination and robustness of the clock". Proceedings of the National Academy of Sciences of the United States of America. 110 (12): 4750–5. doi:10.1073/pnas.1302560110. PMC 3606995. PMID 23471982.
19. Huber AL, Papp SJ, Chan AB, Henriksson E, Jordan SD, Kriebs A, Nguyen M, Wallace M, Li Z, Metallo CM, Lamia KA (November 2016). "CRY2 and FBXL3 Cooperatively Degrade c-MYC". Molecular Cell. 64 (4): 774–789. doi:10.1016/j.molcel.2016.10.012. PMC 5123859. PMID 27840026.
20. Xing W, Busino L, Hinds TR, Marionni ST, Saifee NH, Bush MF, Pagano M, Zheng N (April 2013). "SCF(FBXL3) ubiquitin ligase targets cryptochromes at their cofactor pocket". Nature. 496 (7443): 64–8. doi:10.1038/nature11964. PMC 3618506. PMID 23503662.