Convergent Evolution of Hemoglobin Function in High-Altitude Andean Waterfowl Involves Limited Parallelism at the Molecular Sequence Level

Abstract

A fundamental question in evolutionary genetics concerns the extent to which adaptive phenotypic convergence is attributable to convergent or parallel changes at the molecular sequence level. Here we report a comparative analysis of hemoglobin (Hb) function in eight phylogenetically replicated pairs of high- and low-altitude waterfowl taxa to test for convergence in the oxygenation properties of Hb, and to assess the extent to which convergence in biochemical phenotype is attributable to repeated amino acid replacements. Functional experiments on native Hb variants and protein engineering experiments based on site-directed mutagenesis revealed the phenotypic effects of specific amino acid replacements that were responsible for convergent increases in Hb-O2 affinity in multiple high-altitude taxa. In six of the eight taxon pairs, high-altitude taxa evolved derived increases in Hb-O2 affinity that were caused by a combination of unique replacements, parallel replacements (involving identical-by-state variants with independent mutational origins in different lineages), and collateral replacements (involving shared, identical-by-descent variants derived via introgressive hybridization). In genome scans of nucleotide differentiation involving high- and low-altitude populations of three separate species, function-altering amino acid polymorphisms in the globin genes emerged as highly significant outliers, providing independent evidence for adaptive divergence in Hb function. The experimental results demonstrate that convergent changes in protein function can occur through multiple historical paths, and can involve multiple possible mutations. Most cases of convergence in Hb function did not involve parallel substitutions and most parallel substitutions did not affect Hb-O2 affinity, indicating that the repeatability of phenotypic evolution does not require parallelism at the molecular level.

Mitochondrial physiology in the skeletal and cardiac muscles is altered in torrent ducks, Merganetta armata, from high altitudes in the Andes

“…It is possible that hypoxia exposure associated with breath-holding diving led to exaptations ( pre-adaptations) for life at high altitudes in lowland torrent ducks, and reduced the selective advantage for adaptive changes in some traits in the high-altitude population. Consistent with this idea, both high-and low-altitude populations of torrent ducks have hemoglobin with an affinity (P 50 =33-35 mmHg) that is much greater than most birds (in which P 50 typically ranges between 44 and 52 mmHg) and is comparable to that of the emperor penguin (P 50 =28-36 mmHg) (Christensen and Dill, 1935;Hirsowitz et al, 1977;Lutz, 1980;Wastl and Leiner, 1931;Meir and Ponganis, 2009;Tamburrini et al, 1994;Natarajan et al, 2015). However, unlike highland populations of many other South American waterfowl, highland torrent ducks lack further evolved specialization in hemoglobin sequence or O 2 -binding affinity compared to their lowland counterparts (McCracken et al, 2009a,b).…”

“…However, there is very little genetic variation across altitudinal gradients in torrent ducks and gene flow between high and low altitudes is likely significant (McCracken et al, 2009a,b;Natarajan et al, 2015;K.G.M., unpublished). Therefore, if the observed differences in muscle physiology in highland torrent ducks arose as evolutionary adaptations to the high-altitude environment, then these differences are presumably under very strong selection.…”

“…Putative adaptations seem to have arisen across the O 2 transport pathway that enhance breathing, pulmonary gas exchange, hemoglobin-O 2 affinity, capillarity of the flight muscle and heart, and oxidative capacity of the flight muscle (Petschow et al, 1977;Weibel, 1984;Jessen et al, 1991;Zhang et al, 1996;Milsom, 2006, 2007;McCracken et al, 2009a,b;Scott et al, 2008Scott et al, , 2009aScott et al, ,b, 2015aNatarajan et al, 2015). Much less is known about whether high-altitude species have similarly altered the phenotypes of other muscles to help support other forms of locomotion and/or shivering in the hypoxic and cold environment at high elevation.…”

Breaking RAD: An evaluation of the utility of restriction site associated DNA sequencing for genome scans of adaptation

“…For example, RADseq is very useful for studies of local adaptation where candidate genes have already been identified because RAD markers can be used to generate a null distribution of loci across the genome (Luikart et al 2003). Natarajan et al (2015) recently conducted an excellent study of this sort with candidate alpha-and betaglobin genes in ducks. They compared allele frequencies of these candidate genes to a null distribution generated by RADseq data for high-and low-altitude population pairs of three Andean duck species.…”

Hemoglobin–oxygen affinity in high-altitude vertebrates: is there evidence for an adaptive trend?

“…For example, birds typically express two main Hb isoforms in definitive red blood cells -a major isoform, HbA, which incorporates α-type products of the α A -globin gene, and a minor isoform, HbD, which incorporates α-type products of the α D -globin gene; the two isoforms share the same β-type subunits Grispo et al, 2012;Opazo et al, 2015). In all bird species that have been examined to date, the minor HbD isoform has an appreciably higher O 2 affinity than the major HbA isoform in the presence of allosteric effectors (Grispo et al, 2012;Projecto-Garcia et al, 2013;Cheviron et al, 2014;Galen et al, 2015;Natarajan et al, 2015bNatarajan et al, , 2016. Other sauropsid taxa also tend to co-express multiple, structurally distinct Hb isoforms in definitive erythrocytes (Weber and Jensen, 1988;Storz et al, 2011bStorz et al, , 2015bDamsgaard et al, 2013).…”