The reverse flow of genetic information (the reverse transcription of RNA to DNA) allows retroviruses and retrotransposons to invade our genome. Together, retroelements make up more than 40% of the human genome. The HUSH epigenetic repressor complex defends the genome from retroelement invasion through the recognition and silencing of ‘intronless’ DNA, the hallmark of reverse transcription. Therefore, intronless cDNA provides the ‘abnormal molecular pattern’ which allows HUSH to distinguish foreign retroelements from host genes, making HUSH a part of the innate immune system. This presents a paradox: intronless genes (ILGs) comprise ~3-5% of the human protein-coding genome and are well expressed I.e. resistant to hush silencing despite themselves arising through retroelement invasion. I wanted to understand this paradox and determine how ILGs escape HUSH silencing.

I initially assessed genome-wide RNA binding and transcriptional repression by HUSH to identify nucleic acid sequence determinants of HUSH-sensitivity. My findings suggest that ILGs evolve GC-rich sequence within their amino acid sequence constraints, through GC-biased codon usage and increased 5’UTR GC content. This enables their escape from HUSH despite the absence of introns, allowing beneficial retroelements to be domesticated. Decreasing GC content within an intronless coding sequence incrementally impaired recruitment of TREX, an essential mRNA export complex with a known GC binding RNA preference. This decreased GC content corresponded with reduced RNA export and increased HUSH repression. By tethering TREX components to intronless RNA, I could promote HUSH escape and this was most striking for ALYREF, an essential RNA- binding subunit of TREX which facilitates mRNA packaging. In contrast, depletion of TREX components (THOC1/2) induced HUSH-sensitivity of intronless reporters. As RNA binding by ALYREF and periphilin, the main RNA binding component of HUSH, is inversely proportional, I propose a model in which periphilin and ALYREF compete for nascent RNA binding. My results suggest this binding competition between periphilin and ALYREF determines the balance between HUSH-dependent silencing versus RNA nuclear export and gene expression.