Alternative titles; symbols
HGNC Approved Gene Symbol: RNU6-1
Cytogenetic location: 15q23 Genomic coordinates (GRCh38): 15:67,839,940-67,840,045 (from NCBI)
Kandels-Lewis and Seraphin (1993) examined the role of U6 snRNA in 5-prime splice site selection. Studying the spliceosome in Saccharomyces cerevisiae, Lesser and Guthrie (1993) demonstrated that mutations in U6 snRNA alter splice site specificity.
Ganot et al. (1999) demonstrated that recognition of the correct 2-prime-O-methylation and pseudouridylation sites in U6 snRNA relied on short nucleotide sequences around the target sites. They showed that trans-acting factors directing modification of U6 snRNA at all 2-prime-O-methylation and pseudouridylation sites, such as SNORD6 (618943), were present and functionally active in nucleolus.
The human snRNA promoters contain a proximal sequence element (PSE) required for basal transcription and a distal sequence element (DSE) required for activated transcription. The PSE recruits the multisubunit factor SNAPC (see 605076), whereas the DSE recruits OCT1 (164175). OCT1 and SNAPC bind cooperatively to DNA when their respective binding sites are moved into proximity through a mechanism that involves a defined protein-protein contact. Zhao et al. (2001) showed that on the natural U6 promoter, cooperative binding of OCT1 and SNAPC is mediated by a positioned nucleosome that resides between the DSE and the PSE. This cooperative binding requires the same protein-protein contact as cooperative binding to closely spaced sites on naked DNA and mediates transcription activation.
Valadkhan and Manley (2001) demonstrated that a protein-free complex of 2 snRNAs, U2 (180690) and U6, can bind and position a small RNA containing the sequence of the intron branch site, and activate the branch adenosine to attack a catalytically critical domain of U6 in a reaction that is related to the first step of splicing. Valadkhan and Manley (2001) concluded that their data provide direct evidence for the catalytic potential of spliceosomal snRNAs.
U6 snRNA is posttranscriptionally oligouridynylated by TUT1 (610641), and the last nucleotide of approximately 90% of U6 snRNA is modified to form a terminal 2-prime,3-prime-cyclic phosphate that is required for stability. Using a genetic screen, Mroczek et al. (2012) found that human USB1 complemented loss of Usb1 in yeast, with restored cell growth and splicing of a test pre-mRNA. Addition of U6 snRNA also partially rescued Usb1 deficiency in yeast. Knockdown of USB1 in HeLa cells via small interfering RNA did not alter U6 snRNA levels or significantly affect pre-mRNA splicing, but U6 snRNA molecules became extended with more heterogeneous length compared with controls due to TUT1 3-prime uridylation. Loss of USB1 also modestly decreased U6 snRNA stability. Mroczek et al. (2012) concluded that USB1 catalyzes cleavage of the P-O(5-prime) bond at the 3-prime end of poly(U) in U6 snRNA, resulting in formation of a 2-prime,3-prime-cyclic phosphate and removal of the terminal uridine.
Hilcenko et al. (2013) showed that USB1 progressively trimmed 3-prime poly(A) as well as poly(U) in U6 snRNA and generated a 2-prime,3-prime-cyclic phosphate at the 3-prime end of U6 snRNA. USB1 read nucleotide A102 of U6 snRNA as a pause signal and stopped trimming 5 uridines downstream. In lymphoblasts from patients with poikiloderma with neutropenia (PN; 604173) due to USB1 mutation, Hilcenko et al. (2013) found that U6 snRNA had aberrant nontemplated 3-prime oligo(A) tails, which are characteristic of nuclear RNA surveillance targets. They concluded that USB1 functions in posttranscriptional 3-prime end processing that protects U6 snRNA from targeted destruction by the nuclear exosome.
Yean et al. (2000) showed that yeast U6 snRNA specifically binds a divalent metal ion, probably magnesium, that is required for catalysis of the first step of splicing. With magnesium, U6 snRNA with a sulfur substitution for the 2 nonbridging phosphoryl oxygens of nucleotide U80 reconstituted a fully assembled yet catalytically inactive spliceosome. Adding a thiophilic ion such as manganese allowed the first transesterification reaction to occur but not the second. Magnesium competitively inhibited the manganese-rescued reaction, indicating that the metal-binding site at U6/U80 exists in the wildtype spliceosome and that the site changes its metal requirement for activity in the spliceosome. Yean et al. (2000) concluded that the U6 snRNA contributes to pre-mRNA splicing through metal-ion coordination, which is consistent with RNA catalysis by the spliceosome.
Crystal Structure
Juo et al. (2003) reported a 2.95-angstrom resolution crystal structure of the ternary complex containing BRF1 (604902) homology domain II, the conserved region of TBP (600075), and 19 basepairs of U6 promoter DNA. The structure revealed the core interface for assembly of transcription factor IIIB and demonstrated how the loosely packed BRF1 domain achieves remarkable binding specificity with the convex and lateral surfaces of TBP.
Ganot, P., Jady, B. E., Bortolin, M.-L., Darzacq, X., Kiss, T. Nucleolar factors direct the 2-prime-O-ribose methylation and pseudouridylation of U6 spliceosomal RNA. Molec. Cell. Biol. 19: 6906-6917, 1999. [PubMed: 10490628] [Full Text: https://doi.org/10.1128/MCB.19.10.6906]
Hilcenko, C., Simpson, P. J., Finch, A. J., Bowler, F. R., Churcher, M. J., Jin, L., Packman, L. C., Shlien, A., Campbell, P., Kirwan, M., Dokal, I., Warren, A. J. Aberrant 3-prime oligodenylation of spliceosomal U6 small nuclear RNA in poikiloderma with neutropenia. Blood 121: 1028-1038, 2013. [PubMed: 23190533] [Full Text: https://doi.org/10.1182/blood-2012-10-461491]
Juo, Z. S., Kassavetis, G. A., Wang, J., Geiduschek, E. P., Sigler, P. B. Crystal structure of a transcription factor IIIB core interface ternary complex. Nature 422: 534-539, 2003. [PubMed: 12660736] [Full Text: https://doi.org/10.1038/nature01534]
Kandels-Lewis, S., Seraphin, B. Involvement of U6 snRNA in 5-prime splice site selection. Science 262: 2035-2039, 1993. Note: Erratum: Science 263: 739 only, 1994. [PubMed: 8266100] [Full Text: https://doi.org/10.1126/science.8266100]
Lesser, C. F., Guthrie, C. Mutations in U6 snRNA that alter splice site specificity: implications for the active site. Science 262: 1982-1988, 1993. [PubMed: 8266093] [Full Text: https://doi.org/10.1126/science.8266093]
Mroczek, S., Krwawicz, J., Kutner, J., Lazniewski, M., Kucinski, I., Ginalski, K., Dziembowski, A. C16orf57, a gene mutated in poikiloderma with neutropenia, encodes a putative phosphodiesterase responsible for the U6 snRNA 3-prime end modification. Genes Dev. 26: 1911-1925, 2012. [PubMed: 22899009] [Full Text: https://doi.org/10.1101/gad.193169.112]
Valadkhan, S., Manley, J. L. Splicing-related catalysis by protein-free snRNAs. Nature 413: 701-707, 2001. [PubMed: 11607023] [Full Text: https://doi.org/10.1038/35099500]
Yean, S.-L., Wuenschell, G., Termini, J., Lin, R.-J. Metal-ion coordination by U6 small nuclear RNA contributes to catalysis in the spliceosome. Nature 408: 881-884, 2000. [PubMed: 11130730] [Full Text: https://doi.org/10.1038/35048617]
Zhao, X., Pendergrast, P. S., Hernandez, N. A positioned nucleosome on the human U6 promoter allows recruitment of SNAPc by the Oct-1 POU domain. Molec. Cell 7: 539-549, 2001. [PubMed: 11463379] [Full Text: https://doi.org/10.1016/s1097-2765(01)00201-5]