Evolutionary and functional properties of a two-locus beta-globin polymorphism in Indian house mice

Abstract

Electrophoretic surveys of hemoglobin (Hb) polymorphism in house mice from South Asia and the Middle East have revealed that two alternative beta-globin haplotypes, Hbb(d) and Hbb(p), are often present at intermediate frequencies in geographically disparate populations. Both haplotypes harbor two functionally distinct beta-globin paralogs, HBB-T1 (which encodes the beta-chain subunits of the major Hb isoform) and HBB-T2 (which encodes the beta-chains of the minor Hb isoform). The Hbb(d) and Hbb(p) haplotypes share identical HBB-T1 alleles, but products of the alternative HBB-T2 alleles (d(minor) and p(minor)) are distinguished by two amino acid substitutions. To investigate the possible adaptive significance of the Hbb(d)/Hbb(p) polymorphism we conducted a population genetic analysis of the duplicated beta-globin genes of Indian house mice (Mus castaneus) in conjunction with experimental studies of Hb function in inbred strains of mice that carry the alternative Hbb(d) and Hbb(p) haplotypes. The main objectives of this study were (i) to characterize patterns of nucleotide polymorphism and linkage disequilibrium in the duplicated beta-globin genes of M. castaneus, (ii) to test the hypothesis that the Hbb(d) and Hbb(p) haplotypes are maintained as a balanced polymorphism, and (iii) to assess whether allelic differences in the alternative minor Hb isoforms (d(minor) and p(minor)) are associated with different O(2)-binding properties. A multilocus analysis of polymorphism and divergence revealed that levels of diversity at the HBB-T2 gene exceeded neutral expectations, and reconstructed haplotype networks for both beta-globin paralogs revealed extensive allele sharing with several other closely related species of Mus. However, despite this suggestive evidence for balancing selection, O(2)-equilibrium curves revealed no discernible functional differences between red cell lysates containing the d(minor) and p(minor) Hb isoforms. If the d(minor) and p(minor) alleles are maintained as a balanced polymorphism, our results indicate that the associated fitness variance is not directly related to respiratory functions of Hb.

Figure 1.—

Genomic structure of the β-globin gene family of the house mouse (BALB/c strain) on chromosome 7.

Figure 2.—

Rates of decay of gametic linkage disequilibrium within the HBB-T1 and HBB-T2 genes. The red lines show nonlinear regressions of r² against physical distance using a mutation–recombination–drift model (see text for details). Open symbols denote r² estimates for pairs of polymorphic sites in the HBB-T1 gene, and solid symbols denote estimates for pairs of polymorphic sites in the HBB-T2 gene.

Figure 3.—

Median joining network showing relationships among β-globin coding sequences from M. castaneus and four other species of Mus that are known to segregate the Hbb^d and Hbb^p haplotypes (M. domesticus, M. macedonicus, M. musculus, and M. spicilegus). Haplotype networks for the HBB-T1 and HBB-T2 paralogs are shown in A and B, respectively. The size of each circle is proportional to the corresponding haplotype frequency. Inferred intermediate haplotypes are shown as black circles on branches connecting observed haplotypes. Branches between haplotypes indicate one mutational step, and tick marks denote additional steps. Amino acid changes in HBB-T1 sequences are shown in relation to the canonical d_major/p_major sequence from the BALB/c and MSM/s inbred strains, and amino acid changes in HBB-T2 are shown in relation to the canonical d_minor sequence from BALB/c. The three allele classes at HBB-T1 are indicated in A. The black line in B separates haplotypes that are referable to the d_minor and p_minor allele classes.

Figure 4.—

O₂-equilibrium curves of stripped house mouse Hbs at pH 7.40 and 37° in the presence and absence of allosteric cofactors [(Cl⁻), 0.10 m; (NaHEPES), 0.1 m; DPG/Hb tetramer ratio, 2.0; (Heme), 0.16 mm] and in the presence of the met-Hb reductase system (Hayashi et al. 1973). Representative curves for the Hbs from two inbred strains, BALB/c (which is homozygous for Hbb^d) and MSM/s (which is homozygous for Hbb^p), are shown in A and B, respectively. C shows mean values (± SEM) of P₅₀ (n = 3) measured under the above conditions and indicates that the β-chain isoHbs produced by the Hbb^d haplotype and the Hbb^p haplotype do not differ in intrinsic O₂ affinity (as revealed by the comparison of stripped Hbs) or in sensitivity of Hb–O₂ to the presence of allosteric cofactors such as DPG or Cl⁻ ions.