The methyl phosphate capping enzyme Bin3 is a stable component of the fission yeast telomerase holoenzyme

Porat, Jennifer; Me, Baidouri; Grigull, Jörg; Deragon, Jean-Marc; Bayfield, Mark A.

doi:10.1101/2021.04.08.438850

“…To our knowledge, this is the first example of a member of the LARP7 family functioning in a complex with MePCE/Bin3 outside the 7SK complex, indicating that the functional association of these two proteins predates the metazoan invention of the 7SK RNP as a transcriptional and translational regulator. Coincidentally, RIP-Seq experiments with epitope-tagged Bmc1 (Bin3) also identified TER1 as a target leading to an independent characterization of the Bmc1/Pof8 interaction by the Bayfield group 42 .…”

Section: Discussionmentioning

confidence: 99%

A putative cap binding protein and the methyl phosphate capping enzyme Bin3/MePCE function in telomerase biogenesis

Páez-Moscoso

¹

,

Ho

²

,

Pan

³

et al. 2022

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Telomerase reverse transcriptase (TERT) and the noncoding telomerase RNA (TR) subunit constitute the core of telomerase. Additional subunits are required for ribonucleoprotein complex assembly and in some cases remain stably associated with the active holoenzyme. Pof8, a member of the LARP7 protein family is such a constitutive component of telomerase in fission yeast. Using affinity purification of Pof8, we have identified two previously uncharacterized proteins that form a complex with Pof8 and participate in telomerase biogenesis. Both proteins participate in ribonucleoprotein complex assembly and are required for wildtype telomerase activity and telomere length maintenance. One factor we named Thc1 (Telomerase Holoenzyme Component 1) shares structural similarity with the nuclear cap binding complex and the poly-adenosine ribonuclease (PARN), the other is the ortholog of the methyl phosphate capping enzyme (Bin3/MePCE) in metazoans and was named Bmc1 (Bin3/MePCE 1) to reflect its evolutionary roots. Thc1 and Bmc1 function together with Pof8 in recognizing correctly folded telomerase RNA and promoting the recruitment of the Lsm2-8 complex and the catalytic subunit to assemble functional telomerase.

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“…Using Northern blot and RT-qPCR assays, we showed that the levels of various tRNA and miRNAs are not affected by amus mutations (Figure S4A, B, C). The pombe Bmc1 protein has been shown to be important for telomere length homeostasis by interacting with the telomerase RNA substrate (Páez-Moscoso et al 2022;Porat et al 2022), we therefore measured telomeric transcription in amus mutants but did not observe any significant change (Figure S4D). Therefore, loss of Amus seems to specifically affect the steady state level of U6 snRNA.…”

Section: Amus Is a Nuclear Protein That Interacts With And Controls T...mentioning

confidence: 96%

“…Despite these results suggesting MePCE is the capping enzyme for U6, neither the partial loss of MePCE in mammalian cells nor the complete loss of Bin3 in Drosophila affects cellular U6 levels (Jeronimo et al 2007;Singh et al 2011). More recently, two groups carried out independent studies of the Bin3/MePCE homologous Bmc1 protein in S. pombe, an organism that lacks 7SK but depends on U6 for pre-mRNA splicing (Páez-Moscoso et al 2022;Porat et al 2022). Although U6 association with Bmc1 has been established in one study, both groups concluded that the loss of Bmc1 has no effect on U6 level.…”

Section: Introductionmentioning

confidence: 99%

Drosophila Amus and Bin3 methylases functionally replace mammalian MePCE for capping and the stabilization of U6 and 7SK snRNAs

Qiu

¹

,

Wang

²

,

Ying

³

et al. 2023

Preprint

0

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U6 and 7SK snRNAs process a 5 prime cap, believed to be essential for their stability and maintained by mammalian MePCE or Drosophila Bin3 enzymes. Although loss of either protein results in 7SK instability, loss of neither is associated with U6 reduction. Their yeast homolog is also not required for U6 stability, casting further doubts on the function of capping U6. Here we show that the Drosophila Amus protein, homologous to both Bin3 and MePCE, is essential for U6 but not 7SK stability. A full function of Amus is required for Drosophila development, and that rests primarily on its methylase activity. Remarkably, the loss of U6 is rescued by the expression of an Amus-MePCE hybrid protein harboring the methyltransferase domain from MePCE, highlighting the conserved function of the two proteins as the U6 capping enzyme. Our new investigations in human cells establish a dependence of both U6 and 7SK stability on MePCE, resolving a long-standing uncertainty. While uncovering an interesting division of labor of Bin3/MePCE/Amus proteins, we discovered a Bin3-Box domain present only in enzymes associated with 7SK regulation. Targeted mutagenesis in Drosophila confirmed its importance for Bin3 function, revealing a possible conserved element in 7SK but not U6 biology.

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A putative cap binding protein and the methyl phosphate capping enzyme Bin3/MePCE function in telomerase biogenesis

Páez-Moscoso

¹

,

Ho

²

,

Pan

³

et al. 2021

Preprint

2

3

0

View full text Add to dashboard Cite

Telomerase reverse transcriptase (TERT) and the noncoding telomerase RNA (TR) subunit constitute the core of telomerase. Additional subunits are required for ribonucleoprotein complex assembly and in some cases remain stably associated with the active holoenzyme. Pof8, a member of the LARP7 protein family is such a constitutive component of telomerase in fission yeast. Using affinity purification of Pof8, we have identified two previously uncharacterized proteins that form a complex with Pof8 and participate in telomerase biogenesis. Both proteins participate in ribonucleoprotein complex assembly and are required for wildtype telomerase activity and telomere length maintenance. One factor we named Thc1 (Telomerase Holoenzyme Component 1) shares structural similarity with the nuclear cap binding complex and the poly-adenosine ribonuclease (PARN), the other is the ortholog of the methyl phosphate capping enzyme (Bin3/MePCE) in metazoans and was named Bmc1 (Bin3/MePCE 1) to reflect its evolutionary roots. Thc1 and Bmc1 function together with Pof8 in recognizing correctly folded telomerase RNA and promoting the recruitment of the Lsm2-8 complex and the catalytic subunit to assemble functional telomerase.

show abstract

The methyl phosphate capping enzyme Bin3 is a stable component of the fission yeast telomerase holoenzyme

Cited by 3 publications

References 83 publications

A putative cap binding protein and the methyl phosphate capping enzyme Bin3/MePCE function in telomerase biogenesis

A putative cap binding protein and the methyl phosphate capping enzyme Bin3/MePCE function in telomerase biogenesis

Drosophila Amus and Bin3 methylases functionally replace mammalian MePCE for capping and the stabilization of U6 and 7SK snRNAs

A putative cap binding protein and the methyl phosphate capping enzyme Bin3/MePCE function in telomerase biogenesis

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