Oscillating Evolution of a Mammalian Locus with Overlapping Reading Frames: An XLαs/ALEX Relay

Abstract

XLαs and ALEX are structurally unrelated mammalian proteins translated from alternative overlapping reading frames of a single transcript. Not only are they encoded by the same locus, but a specific XLαs/ALEX interaction is essential for G-protein signaling in neuroendocrine cells. A disruption of this interaction leads to abnormal human phenotypes, including mental retardation and growth deficiency. The region of overlap between the two reading frames evolves at a remarkable speed: the divergence between human and mouse ALEX polypeptides makes them virtually unalignable. To trace the evolution of this puzzling locus, we sequenced it in apes, Old World monkeys, and a New World monkey. We show that the overlap between the two reading frames and the physical interaction between the two proteins force the locus to evolve in an unprecedented way. Namely, to maintain two overlapping protein-coding regions the locus is forced to have high GC content, which significantly elevates its intrinsic evolutionary rate. However, the two encoded proteins cannot afford to change too quickly relative to each other as this may impair their interaction and lead to severe physiological consequences. As a result XLαs and ALEX evolve in an oscillating fashion constantly balancing the rates of amino acid replacements. This is the first example of a rapidly evolving locus encoding interacting proteins via overlapping reading frames, with a possible link to the origin of species-specific neurological differences.

Rapid asymmetric evolution of a dual-coding tumor suppressor INK4a/ARF locus contradicts its function

“…5A). If the two reading frames were under similar selective pressure, we would expect dN const to equal dN alt as was previously reported for GNAS1 and XBP1 (20,21). On the contrary, most dual-coding exons showed strongly asymmetrical evolutionary pressures acting on the two reading frames: constitutive frames, on average, evolved considerably slower than the alternative frames (Fig.…”

“…A comparison with other genes reveals an unusually high dN: 100% (8,467/8,467) of genes have significantly lower dN for exon 1␤ and 99.98% (8,465/8,467) for exon2/ARF. We caution, however, that these rates cannot be readily interpreted in the region of overlap (exon 2), because of the rates of evolution in one frame depend on the context in the other (21,22). Even so, this behavior is drastically different from the evolutionary regimes of two other mammalian genes with overlapping reading frames, GNAS1 and XBP1, where both reading frames seem to rapidly coevolve with dN rates of overlapping reading frames being approximately equal (20,21).…”

“…We caution, however, that these rates cannot be readily interpreted in the region of overlap (exon 2), because of the rates of evolution in one frame depend on the context in the other (21,22). Even so, this behavior is drastically different from the evolutionary regimes of two other mammalian genes with overlapping reading frames, GNAS1 and XBP1, where both reading frames seem to rapidly coevolve with dN rates of overlapping reading frames being approximately equal (20,21). Thus, it seems that substitution events within a single stretch of genomic DNA result in discordant selective pressures on the reading frames on the INK4a/ ARF locus, a pattern previously reported only for short terminal overlaps in bacterial and viral genomes (23 INK4a and ARF interact with a number of other proteins and serve as nodes in independent RB and p53 tumor suppression pathways (Fig.…”

Physiological functions of the imprinted Gnas locus and its protein variants Gαs and XLαs in human and mouse

“…This ORF encodes a protein termed Alex, which is conserved, but unrelated to G-proteins (Klemke et al 2001, Nekrutenko et al 2005. Although Alex was detected in PC12 cells and human platelets (Klemke et al 2001, Freson et al 2003), its abundance, expression level and significance in vivo remain unclarified.…”

Length of the ORF, position of the first AUG and the Kozak motif are important factors in potential dual-coding transcripts

“…As a result, they appear and disappear rapidly, and only functional ones were kept and evolved at a slower rate. For example, in GNAS, XLαs and ALEX evolve in an oscillating fashion, constantly balancing the rates of amino-acid replacement [33]. According to our study, the mRNAs of zincfinger proteins could be the most active family of dualcoding transcripts in terms of birth and death rates.…”