Biology subjects

Giulotto, E.

Publications and source records attributed to Giulotto, E..

6 recordsLinked to original sources

bioRxiv2026

Fully Phased Telomere-to-Telomere Assemblies for Thoroughbred Horse and Donkey Haplotypes derived from a Mule Illuminate the Peculiar Evolution of Equid Centromeres

We present telomere-to-telomere genome assemblies of a Thoroughbred horse and a donkey derived from their mule offspring. Now adopted and annotated by NCBI as reference genomes, these assemblies resolve previously inaccessible regions, including satellite arrays, duplications, and telomeres. Equids are known to exhibit an uncoupling between satellite DNA and centromeric function. The completeness of these assemblies enabled annotation of both satellite-based and satellite-free centromeres, as well as non-centromeric satellite loci, revealing notable centromeric plasticity. They also allowed detailed characterization of the variable binding domains of CENP-A--the epigenetic determinant of centromere identity--and CENP-B, whose association with CENP-A, previously considered typical based on a few model organisms, is absent in equids. Comparative analyses of satellite repeats and centromere positions provide new insights into the accelerated karyotypic reshuffling in equid evolution. These assemblies represent foundational resources for equid genomics and support ongoing initiatives such as the Equine Pangenome Project.

molecular biology↗

bioRxiv2025

Unconventional centromere architectures in Tapirus indicus reveal hotspots for satellite-free centromere formation in Perissodactyla

Centromeres, the chromosomal loci responsible for proper segregation during cell division, play a key role in genome evolution and speciation. While centromere function is highly conserved and epigenetically defined by CENP-A, the underlying DNA sequences are among the most rapidly evolving. Although mammalian centromeres are typically associated with satellite DNA, we previously showed that equids carry numerous satellite-free centromeres. In this study, we investigated centromere and karyotype evolution in the endangered Tapirus indicus, a non-equid Perissodactyl with exceptional karyotypic plasticity. Through CENP-A ChIP-seq analysis on the same individual for which a near-gapless diploid genome assembly was available, we identified both canonical satellite-based centromeres and three satellite-free centromeres, emerging from centromere repositioning and representing the first such centromeres described in a non-equid Perissodactyl species. Comparative genomic analysis uncovered evolutionary hotspots for satellite-free centromere formation across Perissodactyla. Finally, analysis of CENP-B binding showed that T. indicus displays uncoupling between CENP-A and CENP-B, a feature previously observed only in equids. These findings reveal that high centromere plasticity is not unique to equids and support a broader model in which centromere plasticity and CENP-B uncoupling contribute to karyotype evolution in mammals.

genomics↗

bioRxiv2025

TERRA transcripts and promoters from telomeric and interstitial sites

The transcription of human telomeres gives rise to a family of long noncoding RNAs, named TERRA. We previously showed that TERRA transcription is driven by CpG island promoters that are composed by stretches of three types of repeats. Using the human genome assembly that was available at that time, putative promoter sequences were localized at several subtelomeres. In this work, using the T2T-CHM13v2.0 human reference genome, we found that 39 out of 46 subtelomeres contain TERRA promoters and grouped them in classes depending on their organization. We then discovered 106 intrachromosomal TERRA-like promoters, adjacent to interstitial telomeric sequences (ITSs) or far away from them. Fortyseven of these promoters are flanked and may regulate the transcription of coding genes, ncRNAs or pseudogenes. Comparative sequence analysis showed that interstitial and subtelomeric promoters belong to a previously undescribed family of segmental duplications deriving from common ancestral sequences. RT-PCR experiments in seven cell lines demonstrated that TERRA transcripts can be synthesized from ITSs. TERRA expression was always low in primary fibroblasts and HeLa cells while highly variable in the other two telomerase positive (HT1080 and HEK293) and in the three telomerase negative ALT cell lines (GM847, U2OS and VA13). The analysis of RNA-seq data from U2OS, HeLa and HEK293 cells showed that 205 ITSs were transcribed in at least one cell lines. The fraction of transcribed ITSs and the level of their transcription increased with the length of the telomeric repeat stretch. Given the large number of transcribed ITSs, we propose that these loci contribute significantly to the production of the TERRA pool.

molecular biology↗

bioRxiv2024

CENP-A/CENP-B uncoupling in the evolutionary reshuffling of centromeres

BackgroundWhile CENP-A is the epigenetic determinant of the centromeric function, the role of CENP-B, the sole centromeric protein binding a specific DNA sequence (CENP-B-box), remains elusive. In the few mammalian species analyzed so far, the CENP-B box is contained in the major satellite repeat that is present at all centromeres. We previously demonstrated that, in the genus Equus, some centromeres lack any satellite repeat. ResultsHere, we show that, in four Equus species, CENP-B is expressed but does not bind the numerous satellite-free and the majority of satellite-based centromeres while it is localized at several ancestral now inactive centromeres. The absence of CENP-B is related to the lack of CENP-B boxes rather than to peculiar features of the protein itself. CENP-B boxes are comprised in a previously undescribed repeat which is not the major satellite bound by CENP-A. Comparative sequence analysis suggests that this satellite was centromeric in the equid ancestor, lost centromeric function during evolution and gave rise to a short CENP-A bound repeat not containing the CENP-B box but being enriched in dyad symmetries. Centromeres lacking CENP-B are functional and recruit normal amounts of the centromeric proteins CENP-A and CENP-C. ConclusionsWe propose that the uncoupling between CENP-B and CENP-A may have played a role in the evolutionary reshuffling of equid centromeres. This study provides new insights into the complexity of centromere organization in a largely biodiverse world where the majority of mammalian species still have to be studied.

genomics↗

bioRxiv2022

Long-read RNA Sequencing Improves the Annotation of the Equine Transcriptome

1A high-quality reference genome assembly, a biobank of diverse equine tissues from the Functional Annotation of the Animal Genome (FAANG) initiative, and incorporation of long-read sequencing technologies, have enabled efforts to build a comprehensive and tissue-specific equine transcriptome. The equine FAANG transcriptome reported here provides up to 45% improvement in transcriptome completeness across tissue types when compared to either RefSeq or Ensembl transcriptomes. This transcriptome also provides major improvements in the identification of alternatively spliced isoforms, novel noncoding genes, and 3 transcription termination site (TTS) annotations. The equine FAANG transcriptome will empower future functional studies of important equine traits while providing future opportunities to identify allele-specific expression and differentially expressed genes across tissues.

genomics↗

bioRxiv2022

Neocentromere formation through Robertsonian fusion and centromere repositioning during the evolution of zebras

Centromeres are epigenetically specified by the histone H3 variant CENP-A and typically associated to highly repetitive satellite DNA. We previously discovered natural satellite-free neocentromeres in Equus caballus and E. asinus. Here, through ChIP-seq with an anti-CENP-A antibody, we found an extraordinarily high number of centromeres lacking satellite DNA in the zebras E. burchelli (15 of 22) and E. grevyi (13 of 23), demonstrating that the absence of satellite DNA at the majority of centromeres is compatible with genome stability and species survival and challenging the role of satellite DNA in centromere function. Nine neocenstromeres are shared between the two species in agreement with their recent separation. We de novo assembled all neocentromeric regions and improved the reference genome of E. burchelli. Sequence analysis of the CENP-A binding domains revealed that they are LINE-1 and AT-rich with four of them showing DNA amplification. In the two zebras, satellite-free centromeres emerged from centromere repositioning or following Robertsonian fusion. In five chromosomes, the centromeric function arose near the fusion points, which are located within regions marked by traces of ancestral pericentromeric sequences. Therefore, besides centromere repositioning, Robertsonian fusions are an important source of satellite-free centromeres during evolution. Finally, in one case, a neocentromere was seeded on an inversion breakpoint. At eleven chromosomes, whose primary constrictions seemed to be associated to satellite repeats by cytogenetic analysis, neocentromeres were instead located near the ancestral inactivated satellite-based centromeres, therefore, the centromeric function has shifted away from a satellite repeat containing locus to a satellite-free new position.

genomics↗