Resurrecting self-cleaving mini-ribozymes from 40-million-year-old LINE-1 elements in human genome

Long Interspersed Nuclear Element (LINE) retrotransposons play an important role in genomic innovation as well as genomic instability in many eukaryotes including human. Random insertions and extinction through mutational inactivation make them perfectly time-stamped "DNA fossils". Here, we investigated the origin of a self-cleaving ribozyme in 5' UTR of LINE-1. We showed that this ribozyme only requires 35 nucleotides for self-cleavage with a simple but previously unknown secondary-structure motif that was determined by deep mutational scanning and covariation analysis. Structure-based homology search revealed the existence of this mini-ribozyme in anthropoids but not in prosimians. In human, the most homologs of this mini-ribozyme were found in lineage L1PA6-10 but essential none in more recent L1PA1-2 or more ancient L1PA13-15. We resurrected mini-ribozymes according to consensus sequences and confirmed that mini-ribozymes were active in L1PA10 and L1PA8 but not in L1PA7 and more recent lineages. The result paints a consistent picture for the emergence of the active ribozyme around 40 million years ago, just before the divergence of the new world monkeys (Platyrrhini) and old-world monkeys (Catarrhini). The ribozyme, however, subsequently went extinct after L1PA7 emerged around 30 million years ago with a deleterious mutation. This work uncovers the rise and fall of the mini-LINE-1 ribozyme recorded in the "DNA fossils" of our own genome. More importantly, this ancient, naturally trans-cleaving ribozyme (after removing the non-functional stem loop) may find its modern usage in bioengineering and RNA-targeting therapeutics.

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Conservation of the sizes of 53 introns and over 100 intronic sequences for the binding of common transcription factors in the human and mouse genes for type II procollagen (COL2A1)

Biochemical Journal ◽

10.1042/bj3080923 ◽

1995 ◽

Vol 308 (3) ◽

pp. 923-929 ◽

Cited By ~ 21

Author(s):

L Ala-Kokko ◽

A P Kvist ◽

M Metsäranta ◽

K I Kivirikko ◽

B de Crombrugghe ◽

...

Keyword(s):

Transcription Factors ◽

Type Ii ◽

Stem Loop ◽

Col2a1 Gene ◽

Conserved Sequences ◽

Consensus Sequences ◽

Rna Transcripts ◽

First Intron ◽

Intron 2 ◽

Human And Mouse

Over 11,000 bp of previously undefined sequences of the human COL2A1 gene were defined. The results made it possible to compare the intron structures of a highly complex gene from man and mouse. Surprisingly, the sizes of the 53 introns of the two genes were highly conserved with a mean difference of 13%. After alignment of the sequences, 69% of the intron sequences were identical. The introns contained consensus sequences for the binding of over 100 different transcription factors that were conserved in the introns of the two genes. The first intron of the gene contained 80 conserved consensus sequences and the remaining 52 introns of the gene contained 106 conserved sequences for the binding of transcription factors. The 5′-end of intron 2 in both genes had a potential for forming a stem loop in RNA transcripts.

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De novo Sequencing of Novel Mycoviruses From Fusarium sambucinum: An Attempt on Direct RNA Sequencing of Viral dsRNAs

Frontiers in Microbiology ◽

10.3389/fmicb.2021.641484 ◽

2021 ◽

Vol 12 ◽

Author(s):

Yukiyoshi Mizutani ◽

Kazuma Uesaka ◽

Ayane Ota ◽

Matteo Calassanzio ◽

Claudio Ratti ◽

...

Keyword(s):

Rna Sequencing ◽

Reading Ability ◽

High Throughput Sequencing ◽

De Novo ◽

Genomic Sequence ◽

Homology Search ◽

Fusarium Sambucinum ◽

Consensus Sequences ◽

Molecular Features ◽

Wide Range

An increasing number of viruses are continuously being found in a wide range of organisms, including fungi. Recent studies have revealed a wide viral diversity in microbes and a potential importance of these viruses in the natural environment. Although virus exploration has been accelerated by short-read, high-throughput sequencing (HTS), and viral de novo sequencing is still challenging because of several biological/molecular features such as micro-diversity and secondary structure of RNA genomes. This study conducted de novo sequencing of multiple double-stranded (ds) RNA (dsRNA) elements that were obtained from fungal viruses infecting two Fusarium sambucinum strains, FA1837 and FA2242, using conventional HTS and long-read direct RNA sequencing (DRS). De novo assembly of the read data from both technologies generated near-entire genomic sequence of the viruses, and the sequence homology search and phylogenetic analysis suggested that these represented novel species of the Hypoviridae, Totiviridae, and Mitoviridae families. However, the DRS-based consensus sequences contained numerous indel errors that differed from the HTS consensus sequences, and these errors hampered accurate open reading frame (ORF) prediction. Although with its present performance, the use of DRS is premature to determine viral genome sequences, the DRS-mediated sequencing shows great potential as a user-friendly platform for a one-shot, whole-genome sequencing of RNA viruses due to its long-reading ability and relative structure-tolerant nature.

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Regulation of Alpha Hemoglobin Stabilizing Protein (AHSP) mRNA Stability by a 3′ UTR Iron Response Element.

Blood ◽

10.1182/blood.v108.11.535.535 ◽

2006 ◽

Vol 108 (11) ◽

pp. 535-535

Author(s):

Camila O. dos Santos ◽

Louis C. Dore ◽

Sara T. Ollala ◽

Fernando F. Costa ◽

Mitchell J. Weiss

Keyword(s):

Iron Overload ◽

Mrna Stability ◽

Iron Homeostasis ◽

Iron Status ◽

Beta Thalassemia ◽

Mrna Levels ◽

Erythroid Cells ◽

Stem Loop ◽

Consensus Sequences ◽

Alpha Globin

Abstract AHSP is an erythroid protein that binds the alpha globin subunit of hemoglobin (Hb) to maintain its structure and limit its oxidative activity. Biochemical and genetic studies demonstrate that AHSP is essential for normal Hb homeostasis and may act via at least two mechanisms: First, as a molecular chaperone to promote the folding and stability of alpha globin prior to its incorporation into HbA (alpha2beta2). Second, to bind and detoxify excess alpha globin that accumulates in normal erythroid precursors, and to a larger extent, in beta thalassemic ones. The existence of a potential iron responsive element (IRE) in the 3′-UTR of the AHSP mRNA raises the interesting possibility that iron homeostasis impacts on alpha globin stability via AHSP. IREs form stem-loop structures that bind cytosolic iron-sensing proteins (IRPs). Typically, IRP binding to the 3′ UTR stabilizes mRNA. Association with iron releases the IRP, enhancing mRNA degradation. Computational algorithms identified IRE-like stem-loop structures in AHSP mRNA of multiple species, yet the primary sequences deviate significantly from canonical IRE consensus sequences determined by studies of classical IREs, such as Transferrin receptor and Ferritin. Several lines of evidence now show that the AHSP IRE binds IRPs to regulate mRNA stability in an iron-dependent fashion: 1) in vitro gelshift competition assays demonstrate that human and mouse AHSP IREs bind tightly to IRPs. 2) AHSP mRNA co-immunoprecipitates with IRPs in erythroid cells in a fashion that depends on an intact IRE. 3) AHSP mRNA is destabilized by iron in both erythroid and heterologous cells; disruption of the IRE renders the mRNA constitutively unstable. To study how iron regulates AHSP expression in vivo, we treated mice with iron dextran for 10 days and then examined AHSP mRNA in Ter119+ erythroid progenitors by RT-PCR. We found that short-term iron overload reduced AHSP mRNA levels by about 50% (p > 0.05). Our findings indicate that AHSP mRNA stability is regulated by iron via an atypical 3′-UTR IRE. These findings extend the potential repertoire for functional IREs that do not conform to previously defined canonical consensus sequences. In addition, they provide a potential mechanism by which erythroid cells can regulate globin stability according to iron status. For example, induction of AHSP during iron deficiency might stabilize apo- alpha globin that could accumulate from lack of heme. Conversely, iron overload could destabilize alpha globin by reducing AHSP levels. As such, iron overload, which occurs in patients with beta thalassemia, might aggravate the disease by further elevating the levels of toxic free alpha globin.

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