Furano, A. V. | bioRxiv Science

bioRxiv2023

CO-EXPRESSION OF DISTINCT COILED COILS CAN LEAD TO THEIR ENTANGLEMENT

L1 (LINE1) non-LTR retrotransposons are ubiquitous genomic parasites and the dominant transposable element in humans having generated about 40% of their genomic DNA during their [~]100 million years (Myr) of activity in primates. L1 replicates in early embryos, causing genetic diversity and defects, but can be active in some somatic stem cells, tumors and during aging. L1 encodes two proteins essential for retrotransposition: ORF2, a replicase, and ORF1, a coiled coil mediated homo trimer, which functions as a nucleic acid chaperone. Both proteins contain highly conserved domains and preferentially bind their encoding transcript to form an L1 ribonucleoprotein (RNP), which mediates retrotransposition. However, the coiled coil has undergone episodes of substantial amino acid replacement to the extent that a given L1 family can concurrently express multiple ORF1s that differ in the sequence of their coiled coils. Here we show that such distinct ORF1p sequences can become entangled forming heterotrimers when co-expressed from separate vectors.

molecular biology↗

bioRxiv2019

The effect of APOBEC3B deaminase on double-stranded DNA

Mutations mediated by the APOBEC3 (A3) family of single-strand specific cytosine deaminases can accumulate in various cancers, as strand-coordinated clusters and isolated lesions. A3-mediated mutations also occur during normal development, accounting for ~20% of heritable mutations. A3B is an archetypical member of this family and is thought to contribute to both cancer initiation and progression. A3B has a strong preference for C in a TC context and catalyzes hydrolysis of the primary amine of un-paired C to generate U. Subsequent repair generates a distinctive pattern of C-substitutions, which along with their context signify their A3B origin. Although single-stranded DNA is the preferred A3B substrate, we report here that in some instances A3B can deaminate the C of TC in a double-stranded DNA context in vitro. These include C paired to O6-methylguanine (O6meG), to an abasic (AP) site, or to a G adjacent to an AP site. AP sites are the most common lesion in DNA, and O6meG levels increase under alkylating conditions caused by environmental nitrosamines and some chemotherapeutic agents. We also show that elevated expression of A3B can enhance double-stranded breaks induced by the alkylating agent MNNG in mammalian cells, but this effect does not require A3B deaminase activity.

biochemistry↗