* 613372

UFM1-SPECIFIC LIGASE 1; UFL1


Alternative titles; symbols

NOVEL LZAP-BINDING PROTEIN; NLBP
KIAA0776
RCAD


HGNC Approved Gene Symbol: UFL1

Cytogenetic location: 6q16.1     Genomic coordinates (GRCh38): 6:96,521,806-96,555,276 (from NCBI)


TEXT

Cloning and Expression

By sequencing clones obtained from a size-fractionated brain cDNA library, Nagase et al. (1998) cloned KIAA0776, which encodes a deduced 799-amino acid protein. RT-PCR ELISA detected highest expression in ovary and kidney. Lower expression was detected in heart, brain, lung, liver, skeletal muscle, pancreas, and testis, with little to no expression in spleen.

Kwon et al. (2010) reported that full-length KIAA0776 contains 794 amino acids.

Using Northern blot analysis, Wu et al. (2010) found variable expression of 2 Rcad transcripts in all mouse tissues examined. Both were predicted to encode the same 793-amino acid protein. EST database analysis suggested the presence of several splice variants of human RCAD, some of which may encode truncated proteins. Full-length human RCAD had an apparent molecular mass of 90 kD by SDS-PAGE.


Gene Function

Using affinity purification and mass spectrometry, Kwon et al. (2010) identified NLBP as a protein that interacted with LZAP (CDK5RAP3; 608202) in transfected HEK293 cells. They confirmed the interaction by in vitro protein pull-down assays. Domain analysis revealed that an N-terminal region of NLBP and a central region of LZAP were required for the interaction. Knockdown of either NLBP or LZAP via small interfering RNA (siRNA) increased expression of MMP9 (120361) and the invasiveness of U2OS cells. Western blot analysis detected reduced levels of LZAP and NLBP in several invasive hepatocellular carcinoma cell lines, but not in a noninvasive cell line. NLBP inhibited the NF-kappa-B (see 164011) signaling pathway. NLBP and LZAP colocalized in the cytoplasm. Both proteins were subject to ubiquitination, and they appeared to stabilize each other against ubiquitination and degradation via the proteasome.

By immunoprecipitation analysis, Wu et al. (2010) found that epitope-tagged C53 (CDK5RAP3; 608202) interacted with DDRGK1 (616177) and RCAD in HEK293 cells. Size-exclusion chromatography revealed that RCAD and C53 fractionated with protein complexes of around 550 kD. Knockdown of RCAD in HeLa and U2OS cells via siRNA caused a dramatic reduction in C53 and DDRGK1 protein levels, apparently due to ubiquitin-mediated degradation. In contrast, overexpression of RCAD inhibited ubiquitination of these proteins. Knockdown of either RCAD or C53 increased both basal and TNF-alpha (TNFA; 191160)-induced NF-kappa-B activity.


Mapping

By radiation hybrid analysis, Nagase et al. (1998) mapped the KIAA0776 gene to chromosome 6. Kwon et al. (2010) stated that the KIAA0776 gene maps to chromosome 6q16.1.

Zhang et al. (2015) stated that the mouse Ufl1 gene maps to chromosome 4.


Animal Model

Zhang et al. (2015) found that homozygous deletion of Rcad in mice resulted in embryonic lethality as early as day 10.5. Rcad -/- embryos appeared anemic, were smaller, had smaller livers, and had a high number of abnormal multinucleated erythrocytes compared with wildtype. Rcad inducible conditional knockout mice were normal at birth, but as Rcad expression was depleted, they lost substantial body weight, exhibited severe anemia and pancytopenia, and died. The major indices for both red blood cells and white blood cells were significantly lower in Rcad-deficient mice compared with wildtype. Rcad deficiency in mice impaired erythroid differentiation at multiple developmental stages. Examination of hematopoietic stem cells (HSCs) from Rcad conditional knockout mice revealed that loss of Rcad activated p53 (TP53; 191170), decreased HSC proliferation, and increased HSC death and cell cycle arrest. Further analyses demonstrated that Rcad deficiency elevated endoplasmic reticulum stress, activated the unfolded protein response, impaired autophagic degradation, and led to accumulation of reactive oxygen species and other cellular damage, which likely induced DNA damage response, abnormal p53 activation, and HSC death.


REFERENCES

  1. Kwon, J., Cho, H. J., Han, S. H., No, J. G., Kwon, J. Y., Kim, H. A novel LZAP-binding protein, NLBP, inhibits cell invasion. J. Biol. Chem. 285: 12232-12240, 2010. [PubMed: 20164180, images, related citations] [Full Text]

  2. Nagase, T., Ishikawa, K., Suyama, M., Kikuno, R., Miyajima, N., Tanaka, A., Kotani, H., Nomura, N., Ohara, O. Prediction of the coding sequences of unidentified human genes. XI. The complete sequences of 100 new cDNA clones from brain which code for large proteins in vitro. DNA Res. 5: 277-286, 1998. [PubMed: 9872452, related citations] [Full Text]

  3. Wu, J., Lei, G., Mei, M., Tang, Y., Li, H. A novel C53/LZAP-interacting protein regulates stability of C53/LZAP and DDRGK domain-containing protein 1 (DDRGK1) and modulates NF-kappa-B signaling. J. Biol. Chem. 285: 15126-15136, 2010. [PubMed: 20228063, images, related citations] [Full Text]

  4. Zhang, M., Zhu, X., Zhang, Y., Cai, Y., Chen, J., Sivaprakasam, S., Gurav, A., Pi, W., Makala, L., Wu, J., Pace, B., Tuan-Lo, D., Ganapathy, V., Singh, N., Li, H. RCAD/Ufl1, a Ufm1 E3 ligase, is essential for hematopoietic stem cell function and murine hematopoiesis. Cell Death Diff. 22: 1922-1934, 2015. [PubMed: 25952549, related citations] [Full Text]


Bao Lige - updated : 03/14/2019
Patricia A. Hartz - updated : 1/9/2015
Creation Date:
Patricia A. Hartz : 4/20/2010
alopez : 02/23/2021
mgross : 03/14/2019
mgross : 03/14/2019
mgross : 01/16/2015
mcolton : 1/9/2015
mgross : 4/20/2010

* 613372

UFM1-SPECIFIC LIGASE 1; UFL1


Alternative titles; symbols

NOVEL LZAP-BINDING PROTEIN; NLBP
KIAA0776
RCAD


HGNC Approved Gene Symbol: UFL1

Cytogenetic location: 6q16.1     Genomic coordinates (GRCh38): 6:96,521,806-96,555,276 (from NCBI)


TEXT

Cloning and Expression

By sequencing clones obtained from a size-fractionated brain cDNA library, Nagase et al. (1998) cloned KIAA0776, which encodes a deduced 799-amino acid protein. RT-PCR ELISA detected highest expression in ovary and kidney. Lower expression was detected in heart, brain, lung, liver, skeletal muscle, pancreas, and testis, with little to no expression in spleen.

Kwon et al. (2010) reported that full-length KIAA0776 contains 794 amino acids.

Using Northern blot analysis, Wu et al. (2010) found variable expression of 2 Rcad transcripts in all mouse tissues examined. Both were predicted to encode the same 793-amino acid protein. EST database analysis suggested the presence of several splice variants of human RCAD, some of which may encode truncated proteins. Full-length human RCAD had an apparent molecular mass of 90 kD by SDS-PAGE.


Gene Function

Using affinity purification and mass spectrometry, Kwon et al. (2010) identified NLBP as a protein that interacted with LZAP (CDK5RAP3; 608202) in transfected HEK293 cells. They confirmed the interaction by in vitro protein pull-down assays. Domain analysis revealed that an N-terminal region of NLBP and a central region of LZAP were required for the interaction. Knockdown of either NLBP or LZAP via small interfering RNA (siRNA) increased expression of MMP9 (120361) and the invasiveness of U2OS cells. Western blot analysis detected reduced levels of LZAP and NLBP in several invasive hepatocellular carcinoma cell lines, but not in a noninvasive cell line. NLBP inhibited the NF-kappa-B (see 164011) signaling pathway. NLBP and LZAP colocalized in the cytoplasm. Both proteins were subject to ubiquitination, and they appeared to stabilize each other against ubiquitination and degradation via the proteasome.

By immunoprecipitation analysis, Wu et al. (2010) found that epitope-tagged C53 (CDK5RAP3; 608202) interacted with DDRGK1 (616177) and RCAD in HEK293 cells. Size-exclusion chromatography revealed that RCAD and C53 fractionated with protein complexes of around 550 kD. Knockdown of RCAD in HeLa and U2OS cells via siRNA caused a dramatic reduction in C53 and DDRGK1 protein levels, apparently due to ubiquitin-mediated degradation. In contrast, overexpression of RCAD inhibited ubiquitination of these proteins. Knockdown of either RCAD or C53 increased both basal and TNF-alpha (TNFA; 191160)-induced NF-kappa-B activity.


Mapping

By radiation hybrid analysis, Nagase et al. (1998) mapped the KIAA0776 gene to chromosome 6. Kwon et al. (2010) stated that the KIAA0776 gene maps to chromosome 6q16.1.

Zhang et al. (2015) stated that the mouse Ufl1 gene maps to chromosome 4.


Animal Model

Zhang et al. (2015) found that homozygous deletion of Rcad in mice resulted in embryonic lethality as early as day 10.5. Rcad -/- embryos appeared anemic, were smaller, had smaller livers, and had a high number of abnormal multinucleated erythrocytes compared with wildtype. Rcad inducible conditional knockout mice were normal at birth, but as Rcad expression was depleted, they lost substantial body weight, exhibited severe anemia and pancytopenia, and died. The major indices for both red blood cells and white blood cells were significantly lower in Rcad-deficient mice compared with wildtype. Rcad deficiency in mice impaired erythroid differentiation at multiple developmental stages. Examination of hematopoietic stem cells (HSCs) from Rcad conditional knockout mice revealed that loss of Rcad activated p53 (TP53; 191170), decreased HSC proliferation, and increased HSC death and cell cycle arrest. Further analyses demonstrated that Rcad deficiency elevated endoplasmic reticulum stress, activated the unfolded protein response, impaired autophagic degradation, and led to accumulation of reactive oxygen species and other cellular damage, which likely induced DNA damage response, abnormal p53 activation, and HSC death.


REFERENCES

  1. Kwon, J., Cho, H. J., Han, S. H., No, J. G., Kwon, J. Y., Kim, H. A novel LZAP-binding protein, NLBP, inhibits cell invasion. J. Biol. Chem. 285: 12232-12240, 2010. [PubMed: 20164180] [Full Text: https://doi.org/10.1074/jbc.M109.065920]

  2. Nagase, T., Ishikawa, K., Suyama, M., Kikuno, R., Miyajima, N., Tanaka, A., Kotani, H., Nomura, N., Ohara, O. Prediction of the coding sequences of unidentified human genes. XI. The complete sequences of 100 new cDNA clones from brain which code for large proteins in vitro. DNA Res. 5: 277-286, 1998. [PubMed: 9872452] [Full Text: https://doi.org/10.1093/dnares/5.5.277]

  3. Wu, J., Lei, G., Mei, M., Tang, Y., Li, H. A novel C53/LZAP-interacting protein regulates stability of C53/LZAP and DDRGK domain-containing protein 1 (DDRGK1) and modulates NF-kappa-B signaling. J. Biol. Chem. 285: 15126-15136, 2010. [PubMed: 20228063] [Full Text: https://doi.org/10.1074/jbc.M110.110619]

  4. Zhang, M., Zhu, X., Zhang, Y., Cai, Y., Chen, J., Sivaprakasam, S., Gurav, A., Pi, W., Makala, L., Wu, J., Pace, B., Tuan-Lo, D., Ganapathy, V., Singh, N., Li, H. RCAD/Ufl1, a Ufm1 E3 ligase, is essential for hematopoietic stem cell function and murine hematopoiesis. Cell Death Diff. 22: 1922-1934, 2015. [PubMed: 25952549] [Full Text: https://doi.org/10.1038/cdd.2015.51]


Contributors:
Bao Lige - updated : 03/14/2019
Patricia A. Hartz - updated : 1/9/2015

Creation Date:
Patricia A. Hartz : 4/20/2010

Edit History:
alopez : 02/23/2021
mgross : 03/14/2019
mgross : 03/14/2019
mgross : 01/16/2015
mcolton : 1/9/2015
mgross : 4/20/2010