CRISPR RNA

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CRISPR RNA or crRNA is a RNA transcript from the CRISPR locus. [1] CRISPR-Cas (clustered, regularly interspaced short palindromic repeats - CRISPR associated systems) is an adaptive immune system found in bacteria and archaea to protect against mobile genetic elements, like viruses, plasmids, and transposons. [2] The CRISPR locus contains a series of repeats interspaced with unique spacers. These unique spacers can be acquired from MGEs. [2]

Contents

Transcripts of the CRISPR Genetic Locus and Maturation of pre-crRNA. 13 Hegasy CRISPR pre crRNA Wiki E CCBYSA.png
Transcripts of the CRISPR Genetic Locus and Maturation of pre-crRNA.

Pre-crRNA is formed after the transcription of the CRISPR locus and before being processed by Cas proteins. Mature crRNA transcripts contain a partial conserved section of repeat and a sequence of spacer that is complementary to the target DNA. [3] crRNA forms an effector complex with a single nuclease or multiple Cas proteins called a Cascade (CRISPR-associated complex for antiviral defense). [3] [1] Once the effector complex is formed a Cas nuclease or single effector protein will cause interference guided by the crRNA match. [4]

Function

Type-I

Type-I CRISPR systems are characterized by Cas3, a nuclease-helicase protein, and the multi-subunit Cascade (CRISPR-associated complex for antiviral defense). The crRNA can form a complex with the Cas proteins in the Cascade and guide the complex to the target DNA sequence. Cas3 is recruited for the nuclease-helicase activity. [5]

Typically in the Cascade, Cas6 generates the mature crRNAs while Cas5 and Cas7 process and stabilize the crRNA. [6]

Type-II

Type-II CRISPR systems [7] are characterized by the single signature nuclease Cas9. [8] In type-II CRISPR systems crRNA and tracrRNA (trans-activating CRISPR RNA) can form a complex known as the guide RNA or gRNA. [9] The crRNA within the gRNA is what matches up with the target sequence or protospacer after the PAM is found. Once the match is made Cas9 will make a double-stranded break.

Stages of CRISPR immunity for type-I, type-II, and type-III. The Stages of CRISPR immunity.svg
Stages of CRISPR immunity for type-I, type-II, and type-III.

Type-III

Type-III CRISPR systems are characterized by Cas10, an RNA cleaving protein. [10] Similar to type-I, a large subunit effector complex is formed and crRNA guides the complex to the target sequence. Cas6 helps to generate the mature crRNA. [10]

Type-IV

Type-IV CRISPR systems do not have an effector nuclease and are associated with plasmids and prophages. A Cas6-like enzyme is associated with the maturation of the crRNA. Not all type-IV systems have a CRISPR locus and therefore do not have crRNA. [11]

Type-V

Type-V CRISPR systems are characterized by Cas12, a nuclease that can cleave ssDNA, dsDNA, and RNA. [7] Like Cas9, Cas12 is the single effector nuclease. Type-V systems process pre-crRNA without tracrRNA. The mature crRNA in complex with Cas12 target the DNA sequence of interest and cleave the DNA. [12]

Type-VI

Type-VI CRISPR systems are characterized by Cas13, a single effector protein that targets RNA. Like the type-V system, Cas13 can process the pre-crRNA without tracrRNA. The mature crRNA in complex with Cas13 guides the complex to the target RNA and degrades it. [13]

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CRISPR is a family of DNA sequences found in the genomes of prokaryotic organisms such as bacteria and archaea. These sequences are derived from DNA fragments of bacteriophages that had previously infected the prokaryote. They are used to detect and destroy DNA from similar bacteriophages during subsequent infections. Hence these sequences play a key role in the antiviral defense system of prokaryotes and provide a form of acquired immunity. CRISPR is found in approximately 50% of sequenced bacterial genomes and nearly 90% of sequenced archaea.

Guide RNA (gRNA) or singel guide RNA (sgRNA) is a short sequence of RNA that functions as a guide for the Cas9-endonuclease or other Cas-proteins that cut the double-stranded DNA and therebye can be used for gene editing. In bacteria and archaea, gRNAs are a part of the CRISPR-Cas system that serves as an adaptive immune defense that protects the organism from viruses. Here the short gRNAs serve as detectors of foreign DNA and direct the Cas-enzymes that degrades the foreign nucleic acid.

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In molecular biology, trans-activating CRISPR RNA (tracrRNA) is a small trans-encoded RNA. It was first discovered by Emmanuelle Charpentier in her study of the human pathogen Streptococcus pyogenes, a type of bacteria that causes harm to humanity. In bacteria and archaea, CRISPR-Cas constitute an RNA-mediated defense system that protects against viruses and plasmids. This defensive pathway has three steps. First, a copy of the invading nucleic acid is integrated into the CRISPR locus. Next, CRISPR RNAs (crRNAs) are transcribed from this CRISPR locus. The crRNAs are then incorporated into effector complexes, where the crRNA guides the complex to the invading nucleic acid and the Cas proteins degrade this nucleic acid. There are several CRISPR system subtypes.

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Cas9 is a 160 kilodalton protein which plays a vital role in the immunological defense of certain bacteria against DNA viruses and plasmids, and is heavily utilized in genetic engineering applications. Its main function is to cut DNA and thereby alter a cell's genome. The CRISPR-Cas9 genome editing technique was a significant contributor to the Nobel Prize in Chemistry in 2020 being awarded to Emmanuelle Charpentier and Jennifer Doudna.

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<span class="mw-page-title-main">CRISPR interference</span> Genetic perturbation technique

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<span class="mw-page-title-main">Epigenome editing</span>

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<span class="mw-page-title-main">Cas12a</span> DNA-editing technology

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<span class="mw-page-title-main">CRISPR gene editing</span> Gene editing method

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<span class="mw-page-title-main">Anti-CRISPR</span> Group of proteins found in phages

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References

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