Reverse transcriptases prime DNA synthesis.

Area of Science:

• Molecular Biology
• Genetics
• Enzymology

Background:

• Reverse transcriptases (RTs) are enzymes that synthesize DNA from RNA templates, challenging the central dogma.
• While RTs function as DNA polymerases, their relationship to primase-active replicases is distant.
• The de novo primase activity of RTs has not been widely recognized.

Purpose of the Study:

• To investigate the de novo primer synthesis activity of CRISPR-associated RTs (CARTs).
• To determine if this primer synthesis ability is conserved across different RT classes.
• To elucidate the role of RT-dependent priming in biological processes like CRISPR array integration.

Main Methods:

• Biochemical assays to test primer synthesis on RNA and DNA templates.
• Analysis of CRISPR-Cas systems utilizing RT-dependent priming.
• Comparative analysis of RT sequences and activities across diverse classes (group II introns, telomerase, retroviruses).

Main Results:

• CRISPR-associated RTs (CARTs) were identified to directly prime DNA synthesis on both RNA and DNA.
• RT-dependent priming is essential for spacer acquisition and integration in certain CRISPR-Cas systems.
• Primer synthesis activity is conserved in diverse RTs, including group II intron RTs, telomerase, and retroviruses.

Conclusions:

• Reverse transcriptases possess a conserved, innate ability to catalyze de novo DNA primer synthesis.
• This activity is independent of accessory domains or alternative priming mechanisms.
• The findings suggest significant, previously underappreciated roles for RT primer synthesis in various biological pathways.