Biology subjects

Slat, V. A.

Publications and source records attributed to Slat, V. A..

3 recordsLinked to original sources

bioRxiv2026

Dramatically reduced spliceosome, intronome, and splicing efficiency in Cyanidiococcus yangmingshanensis and Cyanidium caldarium

Eukaryotic pre-mRNA splicing is catalyzed by the spliceosome, whose ribonucleoprotein composition and the number of intron substrates it acts upon vary widely across eukaryotic lineages. The red alga Cyanidioschyzon merolae possesses a reduced spliceosome lacking the U1 snRNP, and an unusually small intron repertoire. We asked whether these traits are unique to C. merolae or shared across the related Cyanidiales and Cyanidioschyzonales lineages, as well as how they relate to splicing efficiency under light conditions relevant to photosynthetic growth. Genomic and transcriptomic analysis of C. merolae, Cyanidiococcus yangmingshanensis, and Cyanidium caldarium reveal that all three species harbour a reduced, but broadly conserved, set of splicing proteins. Strikingly, covariance model searches failed to detect U1 snRNA in either C. yangmingshanensis or C. caldarium, establishing U1 loss as a shared feature of all three lineages. We identified only 39 introns in C. merolae, 40 in C. yangmingshanensis, and 54 in C. caldarium. Splicing efficiencies were 42-50%, substantially lower than most organisms in which splicing has been measured, but low splicing is compensated by 2-4x higher expression of intron-containing genes than intron-lacking genes. Notably, light can enhance splicing efficiency in C. merolae and C. yangmingshanensis by up to 100%. Furthermore, the splice site and branch site consensus sequences are highly conserved and similar to those found in hemiascomycetous yeasts such as Saccharomyces cerevisiae. 85% of introns contain an in-frame stop codon with a strong bias towards the 5' end of the intron. These results indicate that dramatic streamlining of the spliceosome and intronome, together with inefficient splicing, predated the divergence of these lineages [~]320 million years ago, and is therefore a defining molecular trait of these extremophilic red algae.

molecular biology↗

bioRxiv2026

Genome Position Does Not Impact Transgene Expression Efficiency in the Ancient Red Alga Cyanidioschyzon merolae

The thermoacidophilic red alga Cyanidioschyzon merolae represents one of the simplest photosynthetic eukaryotes and an ancient divergent group in the primary endosymbiotic Viridiplantae. Because of its [~]16 Mbp genome, containing few introns, and capacity for transgene integration by homologous recombination, it is an emerging chassis for synthetic biology. However, genomic integration sites and scalable transformation methods have not been established to systematically investigate the effect of genome position on transgene expression. Here, we combined bioinformatic genome analysis, liquid-handling robotics, and assays of heterologous protein and metabolite production to establish a reproducible framework for nuclear genome engineering in C. merolae. We mapped and annotated 40 intergenic loci as candidate neutral sites across 16 out of 20 chromosomes and could validate 38 of them through robotic-assisted transformation. Reporter gene expression analysis revealed highly uniform expression at all integration sites across broad populations of transformants, indicating surprising minimal positional effects and transcriptional neutrality. The functional equivalence of these genomic landing pads was determined by expression of a heterologous isoprene synthase, and coupling algal photobioreactors to headspace analysis to quantify isoprene production driven by transgene expression from different integration sites. Single copy transgene integrants, regardless of genome position, exhibited comparable reporter signals and consequent isoprene production. Together, these results provide the first experimentally validated set of neutral integration sites in C. merolae and establish a high-throughput transformation protocol for its genetic engineering in the context of synthetic genome biology.

synthetic biology↗

bioRxiv2023

The fission yeast methylphosphate capping enzyme Bmc1/Bin3 promotes 2'-O- methylation of U6 and pre-mRNA splicing

Splicing requires the tight coordination of dynamic spliceosomal RNAs and proteins. U6 is the only spliceosomal RNA transcribed by RNA Polymerase III and undergoes an extensive maturation process. In humans and fission yeast, this includes addition of a 5 {gamma}-monomethyl phosphate cap by members of the Bin3/MePCE family. Previously, we have shown that the Bin3/MePCE homolog Bmc1 is recruited to the S. pombe telomerase holoenzyme by the LARP7 family protein Pof8, where it acts in a catalytic-independent manner to protect the telomerase RNA and facilitate holoenzyme assembly. Here, we show that Bmc1 and Pof8 also interact in a U6-containing snRNP. We demonstrate that Bmc1 and Pof8 promote 2-O-methylation of U6 and identify and characterize a non-canonical snoRNA that guides this methylation. Further, we show that fission yeast strains deleted of Bmc1 or Pof8 show altered U6 snRNP assembly patterns, supporting a more general role for these factors in guiding noncoding RNP assembly beyond the telomerase RNP. These results are thus consistent with a novel role for Bmc1/MePCE family members in stimulating U6 post-transcriptional modifications.

molecular biology↗