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Enzyme Null Alleles in Natural Populations of Drosophila Melanogaster: Frequencies in a North Carolina Population (Allozymes/Enzyme Deficiencies) ROBERT A

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Enzyme Null Alleles in Natural Populations of Drosophila Melanogaster: Frequencies in a North Carolina Population (Allozymes/Enzyme Deficiencies) ROBERT A Proc. Nati. Acad. Sci. USA Vol. 77, No. 2, pp. 1091-1095, February 1980 Genetics Enzyme null alleles in natural populations of Drosophila melanogaster: Frequencies in a North Carolina population (allozymes/enzyme deficiencies) ROBERT A. VOELKER, CHARLES H. LANGLEY, ANDREW J. LEIGH BROWN*, SEIDO OHNISHI, BARBARA DICKSON, ELIZABETH MONTGOMERY, AND SANDRA C. SMITHt Laboratory of Animal Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709 Communicated by C. Clark Cockerham, November 26,1979 ABSTRACT A Raleigh, NC, population of Drosophila that locus will be underestimated and, concomitantly, the melanogaster was sampled for the presence of enzyme null al- detrimental'effects of nulls on fitness will be overestimated. leles at 25 loci. No nulls were found at any of five X-linked loci. This paper reports data on the frequency of null alleles (see Nulls were recovered at 13 of 20 autosomal loci; the weighted mean frequency for all 20 autosomal loci was estimated to be Methods for definition) at 25 allozyme loci in a Raleigh, NC, 0.0025. A consideration of the effects of these null alleles on population of Drosophila melanogaster. The loci to be screened viability strongly suggests that, although they may contribute were selected on the basis of detectability of nulls by starch gel to so-called polygenic variation, they are not representative of electrophoresis. The two criteria for inclusion of a locus were: the entire genome. (i) the enzyme must be present in sufficiently high concentra- The existence of allozyme polymorphisms in natural popula- tion to be reliably scorable in single fly assays after starch gel tions of most species is now well documented (1). The signifi- electrophoresis; and (ii) because of the technique for detection, cance of allozyme variation to the fitness and evolution of at least two mobility variants were necessary at each locus. Of populations is much less well understood. Whereas in some cases nearly 30 enzymes in category i, only one was eliminated be- there is suggestive evidence that individuals carrying a par- cause no variant allele could be found; because of the necessity ticular allozyme genotype may have greater viabilities and of staining larval or pupal homogenates, which would have fertilities than other genotypes at the locus in question, at the inordinately increased the amount of work involved in vast majority of polymorphic enzyme loci there is no evidence screening, several other (polymorphic) loci were eliminated. that the various electrophoretically detectable alleles are not The remaining 25 loci include ones that are highly polymorphic equivalent in biological function. Of potentially greater im- in natural populations as well as ones that are monomorphic portance to overall fitness are enzyme deficiencies genetically (i.e., no variant alleles had been reported prior to our discovery attributable to so-called "null" alleles. Individuals hemizygous, of them for the construction of our "tester" stocks). Thus, the homozygous, or heterozygous for null alleles at loci coding for sample of loci should not be biased towards detecting nulls at the production of enzymes might be expected to have reduced loci that are polymorphic in natural populations or that are fitnesses, with the depression in fitness being correlated with associated with deleterious effects. the importance of the enzyme function to overall metabo- lism. METHODS Surveys of electrophoretic variation have provided little in- Definition of Enzyme Null Allele. In this report the term formation about the frequencies of null alleles in natural pop- "null" denotes an allele that specifies a product that shows no ulations. In most surveys nulls go undetected, because a null/ catalytic activity as a monomer or homomultimer (depending active allele heterozygote would be indistinguishable from a on the locus) in our in vitro gel staining assay. Such nulls may homozygote for an active allele. However, a few nulls have been produce no protein or a protein that is nonfunctional. For ex- reported in several Drosophila species (1-5), usually at ample, a-glycerolphosphate dehydrogenase functions as an frequencies under 5%. In mammals, esterase nulls have been aggregation of two subunits that are products of a-Gpdh. A reported at appreciable frequencies in Microtus agrestis (6) mutant allele was scored as a null if its product failed to function and in Mus musculus (7). as a mutant homodimer even if there was residual activity in Neel (8) has recently summarized the types of null alleles a heterodimer consisting of that mutant subunit and one normal observed in human populations and their potential importance. subunit. Estimates of frequencies of null alleles in humans have been Detection of Null Alleles. The procedure used for the de- obtained from data on the frequency of individuals affected tection of null alleles is shown in Fig. 1. A chromosome ex- with traits having, a known autosomal or X-linked recessive tracted from a wild fly was made heterozygous with each of two mode of inheritance. However, this method of estimation may mobility variants at the locus in question. Failure of the ex- underestimate the frequency of nulls because only individuals tracted chromosome (allele) to produce a heterozygous phe- affected with pathological conditions seek medical treatment notype when heterozygous with either allele suggested a null and are thereby detected. Consequently, nulls will not be de- allele. Such putative nulls were retested to confirm their failure tected at loci where homozygosity or hemizygosity does not to produce a heterozygous electrophoretic pattern. Stocks of result in pathological conditions. Moreover, if persons exhibiting all confirmed nulls were established. pathological conditions are but a subset of all null hemi- or The overall scheme for scoring the 25 loci is shown in Fig. homozygotes at a given locus, the frequency of null alleles at 2. The symbolism is as follows: B = In(1)FM7,B; Cy = The publication costs of this article were defrayed in part by page * Present address: Imperial Cancer Research Fund, Mill Hill Labo- charge payment. This article must therefore be hereby marked "ad- ratories, Burtonhole Lane, London NW7, United Kingdom.. vertisement" in accordance with 18 U. S. C. §1734 solely to indicate t Present address: Department of Bacteriology and Immunology, this fact. University of North Carolina, Chapel Hill, NC 27514. 1091 Downloaded by guest on September 28, 2021 1092 Genetics: Voelker et al. Proc. Natl. Acad. Sci. USA 77 (1980) SM I ,Cy.t-4GpdhF/c-Gpdhs QQXbwv'/-- Gpdh?d'C Select: e-Gpdh?/'- Gpdh9 '-Gpdh?/SM ,Cy b-GpdhF F If e-Gpdh? is: w-Gpdh Y-Gpdh8 o-GpdhSF i-GpdhnuIl K-Gpdhf C-Gpdh5 .-KGpdhF.-(GpdhS .-tGpdhF-c-GpdhS K-GpdhF K-Gpdhs Expected _ _ Electrophoretic _ __ _ Phenotypes are: __ Origin FIG. 1. Procedure for detection of null alleles. The failure of an extracted allele to produce a heterozygous phenotype with either of two different mobility variants suggests the presence of a null allele. In(2LR)SM1,Cy; btvl = In(2LR)bwvl,bwvl; Ubx = to explain our failure to recover nulls on the X chromosome. If In(3LR)TM6,Ubx; SbSer = In(3LR)TM3,SbSer; Sb = Sb in we assume a frequency of 0.0020 nulls per locus on the X standard sequence. Further details of the balancer chromosomes chromosome, the probability that we would observe 3061 (Fum, and mutant symbols are given in ref. 9. The designation "tester" Hex-A, Gpt, and Zw) alleles and not see any nulls is (1 - denotes a chromosome carrying alleles of electrophoretic mo- 0.0020),'"l = 0.002. Thus, the explanation must lie in their bilities different from those of the respective balancer chro- frequency of occurrence or in their associated deleterious ef- mosome for the loci indicated. The loci screened and their ge- fects. netic map locations are given in Table 1. The map locations are To determine whether the distribution of null alleles over from refs. 10-12 and our unpublished results. The electro- autosomal loci was random, we calculated x2 values for good- phoretic and staining conditions are as in refs. 12-15. The wild ness of fit for a 2 X 20 homogeneity test and for a Poisson dis- males used in the screen were captured in a 2-week period at tribution. Both methods indicated significant interlocus het- the Farmer's Market in Raleigh, NC, during June 1977. All erogeneity in the distribution of null alleles. One possible reason crosses were carried out at room temperature with cornmeal/ for the heterogeneity is differences in mutation rates at the molasses/agar medium. various loci, for which there is suggestive evidence (unpublished results of R. A. Voelker, H. E. Schaffer, and T. Mukai and of RESULTS AND DISCUSSION R. R. Racine, C. H. Langley, and R. A. Voelker). The frequencies of nulls recovered at the 25 loci are presented The autosomal data were examined for correlations between in Table 2. No nulls were recovered at the five X-linked loci. null frequency and (i) estimated allozymic heterozyosity in the Nulls were recovered at 13 of the 20 autosomal loci; the population sampled and (ii) subunit molecular weight. The weighted mean frequencies for all loci and for only the auto- correlation between estimated heterozygosity and null fre- somal loci are 0.0020 and 0.0025, respectively. quency was not significant. The correlation between subunit Our failure to recover any nulls on the X chromosome may molecular weight and null frequency, calculated from molec- be attributable to several factors. First, Pgd is a known lethal ular weight estimates from refs. 17 and 18, unpublished results, as a null hemizygote (16); because wild males (hemizygous) and a personal communication from C.-Y. Lee, was also not were sampled, the recovery of Pgd nulls was precluded. Be- significant.
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