Alternative titles; symbols
HGNC Approved Gene Symbol: BUB1
Cytogenetic location: 2q13 Genomic coordinates (GRCh38): 2:110,637,528-110,678,063 (from NCBI)
| Location | Phenotype |
Phenotype MIM number |
Inheritance |
Phenotype mapping key |
|---|---|---|---|---|
| 2q13 | Colorectal cancer with chromosomal instability, somatic | 114500 | 3 | |
| Microcephaly 30, primary, autosomal recessive | 620183 | Autosomal recessive | 3 |
The BUB1 gene encodes a multifunctional component of the chromosome segregation machinery during mitosis. The protein has an N-terminal kinetochore localization domain, multiple binding motifs, and a C-terminal kinase domain (summary by Carvalhal et al., 2022).
The spindle assembly checkpoint modulates the timing of anaphase initiation in mitotic cells containing improperly aligned chromosomes and increases the probability of successful delivery of a euploid chromosome set to each daughter cell. S. cerevisiae BUB1 is a protein that is required for function of the spindle assembly checkpoint in budding yeast. By searching an EST database using the amino acid sequence of mouse Bub1 as the query, Pangilinan et al. (1997) identified a partial cDNA encoding human BUB1. The 810-amino acid partial human BUB1 protein predicted by this cDNA shows striking sequence conservation with mouse Bub1 along its entire length, as well as significant sequence similarity with S. cerevisiae BUB1. Northern blot analysis detected a 3.8-kb BUB1 transcript in human tissues, with the highest expression in testis and thymus, lower expression in colon and small intestine, and very low expression in spleen and ovary. Northern blot analysis of human BUB1 expression and the expression of another human spindle assembly checkpoint protein, MAD2 (MAD2L1; 601467), revealed a common tissue distribution consistent with roles in a common pathway. In addition, the authors demonstrated that rat Bub1 mRNA accumulates in a rat fibroblast cell transformation system and that the accumulation of rat Bub1 mRNA correlates with the proliferation status of cells in culture.
Using in situ hybridization, Leland et al. (2009) found that Bub1 was expressed ubiquitously in the mouse embryo. In adult mouse tissues, Bub1 expression was highest in thymus, ovary, and testis.
Cahill et al. (1999) reported that the BUB1 gene contains 25 exons. The predicted protein contains a nuclear localization signal in the region between CD1 and CD2.
Cahill et al. (1998) used a genomic clone to map the BUB1 gene to 2q12-q14 by fluorescence in situ hybridization. By analysis of a radiation hybrid panel, Cahill et al. (1999) refined the map position to 2q14. Pangilinan et al. (1997) mapped the mouse Bub1 gene approximately 73 cM from the centromere of chromosome 2.
A key insight leading to the discovery of the molecular basis of microsatellite instability (MIN) in human tumors was the discovery of a similar phenotype in Saccharomyces cerevisiae cells carrying mutations in yeast mismatch repair (MMR) genes. Following this paradigm, Cahill et al. (1998) reasoned that the basis for chromosomal instability (CIN) leading to an abnormal chromosome number (aneuploidy) in human tumor cells might be mitotic checkpoint defects similar to those observed in yeast cells with chromosomal instability. Cells with such defects are expected to exit mitosis prematurely after treatment with microtubule-disrupting agents. To test this hypothesis in human colorectal cancer cells, they treated 4 MIN lines and 6 CIN lines with nocodazole, a microtubule-disrupting drug. As expected, all lines achieved nearly complete cell cycle blocks shortly after this treatment, but a striking morphologic difference was observed between MIN and CIN cells. All MIN cell lines had a normal checkpoint response, resulting in an accumulation of cells with condensed chromosomes characteristic of a sustained mitotic block. In the CIN lines, there was an abnormal response, with many fewer mitotic cells and no clear peak in mitotic index observed at any time point. The response of MIN cells was characteristic of those with intact mitotic checkpoints, and a similar response was observed in normal human fibroblasts. To elucidate the genetic mechanisms underlying the checkpoint defect in CIN cells, Cahill et al. (1998) evaluated the human homolog of S. cerevisiae BUB1. BUB1 is a prototype member of a family of genes, some of which encode proteins that bind to the kinetochore and all of which are required for a normal mitotic delay in response to spindle disruption. They cloned the human homolog of BUB1 and determined the complete coding sequence. Comparison of the yeast and human genes showed that the 2 contain 2 highly conserved domains (CD1 and CD2). CD1 directs kinetochore localization and binding to Bub3, whereas CD2 encodes the kinase domain. In one colorectal cancer cell line, they found a splice site mutation, and in a second cell line, they found a missense mutation in the BUB1 gene.
Lengauer et al. (1998) concluded that most cancers are genetically unstable, but that the instability exists at 2 distinct levels. In a small subset of tumors, the instability is observed at the nucleotide level and results in base substitutions, or in deletions or insertions of a few nucleotides. In most other cancers, the instability is observed at the chromosome level, resulting in losses and gains of whole chromosomes or large portions thereof. They suggested that all chemotherapeutic compounds used at present are more toxic to cancer cells than to normal cells only and specifically because of defective checkpoints that occur in cancer cells. This line of reasoning suggests that, although instability may be essential for neoplasia to develop, it may also prove to be its Achilles' heel when the tumor is attacked by the right agents. Because instabilities reflect defects in cellular processes that maintain the integrity of the genome, they can be expected to generate sensitivities to particular chemical agents. For example, cells with defects in nucleotide-excision repair are sensitive to ultraviolet light, and cells with defective BRCA2 (600185) genes are sensitive to ionizing radiation (Abbott et al., 1998).
Tang et al. (2004) found that depletion of BUB1 and SGO1 (609168) in HeLa cells by RNA interference (RNAi) caused massive missegregation of sister chromatids that originated at centromeres. Loss of chromatid cohesion in BUB1 and SGO1 RNAi cells did not appear to require activation of separase (see 604143), but instead triggered mitotic arrest dependent on MAD2 and Aurora B (604970). Tang et al. (2004) determined that BUB1 maintains the steady-state level and centromeric localization of SGO1, and they concluded that BUB1 protects centromeric cohesion in mitosis through SGO1.
Tang et al. (2004) found that HeLa cells depleted of BUB1 by RNA interference were defective in checkpoint signaling. BUB1 directly phosphorylated CDC20 (603618) in vitro and inhibited the ubiquitin ligase activity of the anaphase-promoting complex (APC), of which CDC20 is a regulatory subunit. A CDC20 mutant lacking all 6 BUB1 phosphorylation sites was refractory to BUB1 mediated phosphorylation and inhibition in vitro. Upon checkpoint activation, BUB1 itself was hyperphosphorylated, and its kinase activity toward CDC20 was stimulated. Ectopic expression of the nonphosphorylatable CDC20 mutant allowed HeLa cells to escape from mitosis in the presence of spindle damage. Tang et al. (2004) concluded that BUB1-mediated phosphorylation of CDC20 is required for proper checkpoint signaling and that inhibition of APC by BUB1 may partly account for the sensitivity of the spindle checkpoint.
Kawashima et al. (2010) demonstrated that BUB1 phosphorylates the conserved ser121 of histone H2A in fission yeast S. pombe. The h2a-SA mutant, in which all cellular H2A-S121 is replaced with alanine, phenocopies the Bub1 kinase-dead mutant (bub1-KD) in losing the centromeric localization of shugoshin proteins (see SGOL1, 609168). Artificial tethering of shugoshin to centromeres largely restores the h2a-SA or bub1-KD-related chromatin instability defects, a function that is evolutionarily conserved. Kawashima et al. (2010) concluded that Bub1 kinase creates a mark for shugoshin localization and the correct partitioning of chromosomes.
Primary Microcephaly 30, Autosomal Recessive
In 2 unrelated patients (P1 and P2) with autosomal recessive primary microcephaly-30 (MCPH30; 620183), Carvalhal et al. (2022) identified homozygous or compound heterozygous mutations in the BUB1 gene (602452.0003-602452.0005). The patients were ascertained through the GeneMatcher program. The mutations, which were found by whole-exome sequencing and confirmed by Sanger sequencing, segregated with the disorder in both families. In vitro studies of patient cells showed multiple mitotic defects in both, although there were partial differences between the 2 patients, likely reflecting differences in residual protein levels or function. Patient fibroblasts showed delays in mitotic progression, variable chromosome segregation defects, and chromosome alignment defects by distinct molecular pathways, including defective BUBR1 localization (P1 cells) or impaired kinase activity (P2 cells). Fibroblasts and lymphocytes from P2 also showed aneuploidy. Cells from both patients showed impaired centromeric localization of TOP2A (126430). The findings illustrated that defects in sister chromatid cohesion and segregation resulting from BUB1 mutations underlie a neurodevelopmental disorder with microcephaly.
Associations Pending Confirmation
De Voer et al. (2013) performed genomewide and targeted copy number and mutation analyses of germline DNA from 208 patients with familial or early-onset (40 years of age or younger) colorectal cancer (CRC; see 114500), identifying haploinsufficiency or heterozygous mutations in the spindle assembly checkpoint genes BUB1 and BUB3 (603719) in 2.9%. In addition to CRC, these patients had variegated aneuploidies in multiple tissues and variable dysmorphic features. De Voer et al. (2013) concluded that these results indicated that mutations in BUB1 and BUB3 cause mosaic variegated aneuploidy and increase the risk of colorectal cancer at a young age. De Voer et al. (2013) identified 3 patients with germline BUB1 mutations. One was a female patient of European ancestry who had a 1.7-Mb deletion, presented with colorectal cancer at the age of 37, and had no family history of cancer. The patient did have mosaic aneuploidy and dysmorphic features, including asymmetric face and hairline as well as a prominent forehead. Her IQ was normal. She had had 2 pregnancies: a spontaneous miscarriage prior to 8 weeks, and the birth of 1 healthy son. The other 2 patients reported were of Han Chinese ancestry. One with a nonsense mutation in BUB1 had multiple GI cancers as well as a renal pelvis carcinoma and lung cancer. He was diagnosed at age 34 years and died at age 47. He had a family history of esophageal cancer, colorectal cancer, and colon polyps. He also had an MLH1 (120436) splice site mutation. The third patient had a BUB1 frameshift mutation and was diagnosed with colon cancer at the age of 31. He had a family history of esophageal cancer, gastric cancer, and colon polyps. He died at age 36. No other cancer susceptibility gene mutations were identified in this patient.
Leland et al. (2009) found that homozygous Bub1 deletion in mice was embryonic lethal. Bub1 +/- mice appeared normal, but Bub1 +/- females were subfertile, and fertility declined further with age. Bub1 +/- males showed normal fertility. Reduced fertility in Bub1 +/- females was associated with impaired preimplantation development, reduced implantation, and arrested embryonic development, irrespective of embryo genotype. Leland et al. (2009) found that these defects were caused by germ cell aneuploidy, with the majority of chromosomal segregation errors occurring during meiosis I.
In a colorectal cancer cell line (V400) with chromosomal instability (see 114500) and a defect in a mitotic checkpoint, Cahill et al. (1998) found a 197-bp deletion, predicted to remove codons 76 to 141 of the BUB1 cDNA and to create a frameshift immediately thereafter. Sequencing of the relevant region of genomic DNA identified a G-to-A transition at the canonical splice donor site, i.e., at the first intronic nucleotide following codon 140.
In a colorectal cancer cell line with chromosomal instability (see 114500) and loss of a mitotic checkpoint (V429), Cahill et al. (1998) identified a missense mutation at codon 492 that resulted in the substitution of tyrosine for a conserved serine. In both V400 (602452.0001) and V429, the mutation was heterozygous and the second allele of the pair was wildtype. Analysis of DNA derived from archived tissues of the patients from whom these cell lines were derived revealed that the mutations were somatic, present in their primary tumors but not in their normal tissues.
In a 3-year-old boy (P1), born of reportedly unrelated Austrian parents, with autosomal recessive primary microcephaly-30 (MCPH30; 620183), Carvalhal et al. (2022) identified a homozygous c.2T-G transversion (c.2T-G, NM_004336.4) in the start codon of the BUB1 gene. The mutation, which was found by whole-exome sequencing and confirmed by Sanger sequencing, was inherited from the unaffected parents, who were each heterozygous. The family was not known to be consanguineous, but the mutation occurred in a region of homozygosity. The variant was found at a low frequency in heterozygous state in the gnomAD database (8.5 x 10(-6)). Patient cells showed normal mRNA levels, but decreased BUB1 protein levels, although low levels of a full-length protein were detectable. There was no evidence of use of an alternative start codon. In vitro studies showed that BUB1 recruitment to kinetochores was nearly undetectable. Kinase activity was partially retained. Patient fibroblasts showed significant delays in mitosis progression with prolonged chromosome alignment and increased segregation defects at mitotic exit compared to controls. There were lagging chromosomes with DNA bridges associated with defective localization of BUBR1 (602860). However, aneuploidy was barely detectable. Finally, patient cells showed impaired centromeric recruitment of TOP2A (126430).
In a 16-year-old girl (P2) with autosomal recessive primary microcephaly-30 (MCPH30; 620183), Carvalhal et al. (2022) identified compound heterozygous mutations in the BUB1 gene: a G-to-A transition in intron 21 (c.2625+1G-A, NM_004336.4) and a 1-bp duplication (c.2197dupG; 602452.0005), presumably resulting in a frameshift and premature termination. The mutations, which were found by whole-exome sequencing and confirmed by Sanger sequencing, were each inherited from an unaffected parent. The splice site mutation had an allele frequency of 4.06 x 10(-6) in the gnomAD database with no homozygotes, whereas the duplication was not present in gnomAD. The splice site mutation was demonstrated to cause the skipping of exon 21, resulting in an in-frame deletion of 54 amino acids in the kinase domain of the BUB1 protein. Patient fibroblasts showed normal mRNA levels, but severely reduced BUB1 protein levels. A shorter protein product corresponding to the kinase-deletion mutant was detected. BUB1 recruitment to kinetochores was reduced compared to controls, and kinase activity was virtually absent. Patient fibroblasts showed significant delays in mitosis progression with prolonged chromosome alignment and increased segregation defects at mitotic exit compared to controls. There were lagging chromosomes with DNA bridges associated with defective localization of BUBR1 (602860). There was also a high frequency of anaphase bridges without lagging centromeres. Patient fibroblasts and lymphocytes showed chromosome number alterations, consistent with aneuploidy. Finally, patient cells showed impaired centromeric recruitment of TOP2A (126430), SGO1 (609168), and AURKB (604970).
For discussion of the 1-bp duplication (c.2197dupG, NM_004336.4) in the BUB1 gene that was found in compound heterozygous state in a patient with autosomal recessive primary microcephaly-30 (MCPH30; 620183) by Carvalhal et al. (2022), see 602452.0004.
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Tang, Z., Sun, Y., Harley, S. E., Zou, H., Yu, H. Human Bub1 protects centromeric sister-chromatid cohesion through Shugoshin during mitosis. Proc. Nat. Acad. Sci. 101: 18012-18017, 2004. [PubMed: 15604152] [Full Text: https://doi.org/10.1073/pnas.0408600102]