Gene Duplication and the Evolution of Hemoglobin Isoform Differentiation in Birds

The majority of bird species co-express two functionally distinct hemoglobin (Hb) isoforms in definitive erythrocytes: HbA (the major adult Hb isoform, with α-chain subunits encoded by the αA-globin gene) and HbD (the minor adult Hb isoform, with α-chain subunits encoded by the αD-globin gene). The αD-globin gene originated via tandem duplication of an embryonic α-like globin gene in the stem lineage of tetrapod vertebrates, which suggests the possibility that functional differentiation between the HbA and HbD isoforms may be attributable to a retained ancestral character state in HbD that harkens back to a primordial, embryonic function. To investigate this possibility and to examine other aspects of the evolution of the avian α-like globin genes, in collaborative effort with the Roy E. Weber lab, Joana Projecto-Garcia, Chandrasekhar Nataraja, and Hideaki Moriyama, we conducted a combined analysis of protein biochemistry and sequence evolution to characterize the structural and functional basis of Hb isoform differentiation in birds. The main objectives were: (1) to characterize the O2-binding properties of HbA and HbD in species that are representative of several major avian lineages; (2) to gain insight into the possible structural basis of the observed functional differentiation between the HbA and HbD isoforms; and (3) to determine whether functional differentiation between the HbA and HbD isoforms is primarily attributable to post-duplication substitutions that occurred in the αA- and αD-globin gene lineages, or whether the differentiation is attributable to substitutions that occurred in the single-copy, pre-duplication ancestor of the αD- and αE-globin genes, in which case the distinctive properties of HbD may represent a retained ancestral character state that is shared with embryonic Hb. Functional experiments involving purified HbA and HbD isofoms from 12 different bird species confirmed that HbD is characterized by a consistently higher O2-affinity in the presence of allosteric effectors such as organic phosphates and Cl- ions. In the case of both HbA and HbD, analyses of oxygenation properties under the two-state Monod-Wyman-Changeux allosteric model revealed that the pH-dependence of Hb-O2 affinity stems from changes in the O2 association constant of deoxy (T-state) Hb. Ancestral sequence reconstructions indicated that the replacement substitutions that distinguish the avian αA- and αD-globin genes occurred exclusively on post-duplication branches of the gene family phylogeny, suggesting that the observed functional differences between the HbA and HbD isoforms are not attributable to the retention of an ancestral (pre-duplication) character state in the αD-globin gene. Adviser: Jay F. Stor

Epigenetic Variation May Compensate for Decreased Genetic Variation with Introductions: A Case Study Using House Sparrows (Passer domesticus) on Two Continents

Epigenetic mechanisms impact several phenotypic traits and may be important for ecology and evolution. The introduced house sparrow (Passer domesticus) exhibits extensive phenotypic variation among and within populations. We screened methylation in populations from Kenya and Florida to determine if methylation varied among populations, varied with introduction history (Kenyan invasion <50 years old, Florida invasion ~150 years old), and could potentially compensate for decrease genetic variation with introductions. While recent literature has speculated on the importance of epigenetic effects for biological invasions, this is the first such study among wild vertebrates. Methylation was more frequent in Nairobi, and outlier loci suggest that populations may be differentiated. Methylation diversity was similar between populations, in spite of known lower genetic diversity in Nairobi, which suggests that epigenetic variation may compensate for decreased genetic diversity as a source of phenotypic variation during introduction. Our results suggest that methylation differences may be common among house sparrows, but research is needed to discern whether methylation impacts phenotypic variation

Gene Duplication and the Evolution of Hemoglobin Isoform Differentiation in Birds

The majority of bird species co-express two functionally distinct hemoglobin (Hb) isoforms in definitive erythrocytes: HbA (the major adult Hb isoform, with α-chain subunits encoded by the αA-globin gene) and HbD (the minor adult Hb isoform, with α-chain subunits encoded by the αD-globin gene). The αD-globin gene originated via tandem duplication of an embryonic α-like globin gene in the stem lineage of tetrapod vertebrates, which suggests the possibility that functional differentiation between the HbA and HbD isoforms may be attributable to a retained ancestral character state in HbD that harkens back to a primordial, embryonic function. To investigate this possibility and to examine other aspects of the evolution of the avian α-like globin genes, in collaborative effort with the Roy E. Weber lab, Joana Projecto-Garcia, Chandrasekhar Nataraja, and Hideaki Moriyama, we conducted a combined analysis of protein biochemistry and sequence evolution to characterize the structural and functional basis of Hb isoform differentiation in birds. The main objectives were: (1) to characterize the O2-binding properties of HbA and HbD in species that are representative of several major avian lineages; (2) to gain insight into the possible structural basis of the observed functional differentiation between the HbA and HbD isoforms; and (3) to determine whether functional differentiation between the HbA and HbD isoforms is primarily attributable to post-duplication substitutions that occurred in the αA- and αD-globin gene lineages, or whether the differentiation is attributable to substitutions that occurred in the single-copy, pre-duplication ancestor of the αD- and αE-globin genes, in which case the distinctive properties of HbD may represent a retained ancestral character state that is shared with embryonic Hb. Functional experiments involving purified HbA and HbD isofoms from 12 different bird species confirmed that HbD is characterized by a consistently higher O2-affinity in the presence of allosteric effectors such as organic phosphates and Cl- ions. In the case of both HbA and HbD, analyses of oxygenation properties under the two-state Monod-Wyman-Changeux allosteric model revealed that the pH-dependence of Hb-O2 affinity stems from changes in the O2 association constant of deoxy (T-state) Hb. Ancestral sequence reconstructions indicated that the replacement substitutions that distinguish the avian αA- and αD-globin genes occurred exclusively on post-duplication branches of the gene family phylogeny, suggesting that the observed functional differences between the HbA and HbD isoforms are not attributable to the retention of an ancestral (pre-duplication) character state in the αD-globin gene. Adviser: Jay F. Stor

Gene Duplication and the Evolution of Hemoglobin Isoform Differentiation in Birds

The majority of bird species co-express two functionally distinct hemoglobin (Hb) isoforms in definitive erythrocytes as follows: HbA (the major adult Hb isoform, with α-chain subunits encoded by the αA-globin gene) and HbD (the minor adult Hb isoform, with α-chain subunits encoded by the αD-globin gene). The αD-globin gene originated via tandem duplication of an embryonic α-like globin gene in the stem lineage of tetrapod vertebrates, which suggests the possibility that functional differentiation between the HbA and HbD isoforms may be attributable to a retained ancestral character state in HbD that harkens back to a primordial, embryonic function. To investigate this possibility, we conducted a combined analysis of protein biochemistry and sequence evolution to characterize the structural and functional basis of Hb isoform differentiation in birds. Functional experiments involving purified HbA and HbD isoforms from 11 different bird species revealed that HbD is characterized by a consistently higher O2 affinity in the presence of allosteric effectors such as organic phosphates and Cl− ions. In the case of both HbA and HbD, analyses of oxygenation properties under the two-state Monod-Wyman-Changeux allosteric model revealed that the pH dependence of Hb-O2 affinity stems primarily from changes in the O2 association constant of deoxy (T-state)-Hb. Ancestral sequence reconstructions revealed that the amino acid substitutions that distinguish the adult-expressed Hb isoforms are not attributable to the retention of an ancestral (pre-duplication) character state in the αD-globin gene that is shared with the embryonic α-like globin gene. Includes supplemental tables & references

Epigenetic Variation May Compensate for Decreased Genetic Variation with Introductions: A Case Study Using House Sparrows (\u3cem\u3ePasser domesticus\u3c/em\u3e) on Two Continents

Epigenetic mechanisms impact several phenotypic traits and may be important for ecology and evolution. The introduced house sparrow (Passer domesticus) exhibits extensive phenotypic variation among and within populations. We screened methylation in populations from Kenya and Florida to determine if methylation varied among populations, varied with introduction history (Kenyan invasion \u3c 50 years old, Florida invasion ~150 years old), and could potentially compensate for decrease genetic variation with introductions. While recent literature has speculated on the importance of epigenetic effects for biological invasions, this is the first such study among wild vertebrates. Methylation was more frequent in Nairobi, and outlier loci suggest that populations may be differentiated. Methylation diversity was similar between populations, in spite of known lower genetic diversity in Nairobi, which suggests that epigenetic variation may compensate for decreased genetic diversity as a source of phenotypic variation during introduction. Our results suggest that methylation differences may be common among house sparrows, but research is needed to discern whether methylation impacts phenotypic variation