Nancy E. Simpson

Genetic Variation in Human Erythrocyte Acetylcholinesterase

A method for solubilization of human erythrocyte membranes was developed and used to survey 70 unselected human blood samples for isozymic variation of stromal acetylcholinesterase. Three variants were observed. Pedigrees of families studied by this method indicated that this variation represented the phenotypic expression of two codominant alleles at a single locus.

Localization of X-Linked Dominant Charcot-Marie-Tooth Disease (CMT 2) to Xq13

A family is described in which Charcot-Marie-Tooth disease is inherited as an X-linked dominant mutation (CMT2). Ten DNA marker loci on the X chromosome were used to map the disease locus by linkage analysis. The DXYS1 sequence at Xq13 was found to be linked to the CMT2 locus at an estimated distance of 6 cM (Zmax = 2.87 at theta max = 0.06). The data also suggested close linkage of the CMT2 locus to PGK1 (Zmax = 1.51 at theta max = 0) which has also been mapped to Xq13. Another DNA locus (DXS3), in the Xq21.3----Xq22 region, did not show close linkage (Zmax = -2.231 at theta max = 0.01). We conclude that the CMT2 locus is probably in or close to band Xq13.

Regional localization of 18 human X-linked DNA sequences

A series of human probes with unique sequences has been isolated from a recombinant phage library constructed with DNA obtained from a human-hamster hybrid cell line. This cell line contained the X chromosome as the only human component. For 18 of these probes, a human X-chromosome origin has been confirmed and they have been regionally assigned by a combination of techniques: dosage studies utilizing DNA from human fibroblasts carrying X-chromosome duplications and deletions; the presence or absence of hybridization to digested DNA from hybrid lines carrying fragments of the X chromosome; and in situ hybridization to metaphase chromosomes. The use of dosage as a means to regionally assign probes significantly improves resolution of the X chromosome.

The β subunit locus of the human fibronectin receptor: DNA restriction fragment length polymorphism and linkage mapping studies

SummaryThe beta subunit of the human fibronectin receptor (FNRB) is a transmembrane protein belonging to the VLA (very late antigens of activation) family. Using pGEM-32, a 2.5-kb partial cDNA clone corresponding to the 3′ portion of the human FNRB locus, multiple restriction fragment length polymorphisms (RFLPs) were revealed on DNAs from unrelated Caucasians. RFLPs detected by five enzymes, BanII, HinfI, KpnI, BglII, and SacI, are of the simple two-allele form, and pairwise linkage analyses of these RFLPs with numerous known DNA markers from the chromosome-10 pericentromeric region not only confirmed the chromosome-10 assignment of the functional FNRB gene but also supported its localization at p11.2 suggested by in situ hybridization. An infrequent MspI RFLP was detected by pB/R2, a 4.6-kb genomic clone from the FNRB locus. Another type of DNA polymorphism was also revealed by the cDNA clone and it was visualized on the Southern blot analyses as the presence or absence of an extra band (or a set of extra bands). It seems to stem from a stretch of DNA sequence present in some individuals at one single locus but absent in others, and is of non-chromosome-10 origin based on linkage analyses with known chromosome 10 markers. This “presence/absence” type of polymorphism could be revealed by all of the 25 restriction enzymes tested and is similar in nature to that previously reported with one of the human dihydrofolate reductase pseudogenes, DHFRP1. Dissection of the pGEM-32 clone demonstrated that the region revealing the non-chromosome-10 sequences is within a fragment about 1.7 kb in length extending from about 600 nucleotides preceding the stop codon down to the end of the cloned FNRB 3′ untranslated region. Due to its high polymorphism information content (PIC) value (0.71 for haplotypes of BanII, HinfI, and KpnI RFLPs) and proximity to the centromere, FNRB will prove to be a highly useful marker for genetic linkage studies of multiple endocrine neoplasia type 2A (MEN2A) as well as for chromosome-10 linkage studies in general.

A linkage group of five DNA markers on human chromosome 10.

Five chromosome 10 DNA markers (D10S1, D10S3, D10S4, D10S5, and RBP3) were typed in five large pedigrees with multiple endocrine neoplasia type 2A (MEN-2A) and in five non-MEN-2A pedigrees. Linkage analyses showed that these loci and the locus for MEN-2A (MEN2A) are in one linkage group spanning at least 70 cM. The order of the marker loci is RBP3-D10S5-D10S3-D10S1-D10S4, with interlocus recombination frequencies of 7, 13-19, 19, and 19%, respectively, all on the same side of MEN2A. Analyses of sex-specific recombination frequencies indicated no significant differences between males and females for any of the map intervals studied. Previous localization of D10S5 and RBP3 to the proximal region of the long arm and the pericentric region, respectively, comparison of results with other studies, and our preliminary results with other chromosome 10 markers suggest that the D10S4 end of the map extends into the long arm. Our linkage map has been constructed using only two- and three-locus analyses. It will be possible to combine our results with those of other groups to construct a more detailed and accurate genetic map of chromosome 10.

Group-Specific Component, Alpha1-Antitrypsin and Esterase D in Canadian Eskimos

Three genetic markers - group-specific component (Gc), alpha1-antitrypsin, and esterase D - were examined in a population of Eskimos from Igloolik in the eastern Canadian Arctic. Gc and esterase D were found to be polymorphic. In addition to the common Gc types, an anodal variant called Gc Igloolik was found, probably identical to previously reported Gc Eskimo. Gene frequencies were Gc1: 0.6524, Gc2: 0.3373, GcIgl: 0.0104, for 338 Eskimos. Genetic types of alpha1-antitrypsin (Pi types) were mostly M, with two MS sibs who were half Caucasian, in 170 Eskimos. Frequencies of the esterase D allele in 336 Eskimos were EsD1: 0.7083, EsD2: 0.2917. The frequencies of Gc2 and EsD2 are both higher than are found in Caucasian populations.

The Genetic Distance between the Coagulation Factor IX Gene and the Locus for the Fragile X Syndrome: Clinical Implications

In 3 families with the fragile-X [fra(X)] syndrome, we have identified a minimum of 4 recombinations in 9 meioses between the syndrome locus and the coagulation Factor IX gene. Two Factor IX intragenic restriction fragment length polymorphisms (RFLPs), produced with TaqI and XmnI, were used as markers. In lod score calculations, incomplete penetrance of the fra(X) mutation in males and females was taken into account by the computer program LIPED. The cumulative maximum lod score calculated from these data and from data previously reported was 2.75 at a recombination frequency of 20% (theta = 0.20). This indicates that the genetic distance between the Factor IX gene and the fra(X) locus is too great for Factor IX probes to be used alone for carrier detection in the fra(X) syndrome. Additional polymorphic loci more tightly linked to the fra(X) syndrome locus are required.

The incidence of type 1 (insulin-dependent) diabetes in Toronto.

SummaryAn incidence of 9.0 out of 100,000 children under 19 years of age with Type 1 (insulin-dependent) diabetes was detected in Toronto during a 2 year prospective study. An increased number of cases occurred in the winter months of one of the years but not of the other during the study. The annual incidence was the same in each year. There were slightly more boys than girls but this was not significant. The diabetic children were significantly younger than their non-affected siblings. The incidence of Type 1 diabetes in Toronto is similar to other North American studies.

GENETICS OF ESTERASES IN MAN

Most of the known genetic enzyme deficiencies and variants in man are rare and have been recognized through the study of metabolic diseases. Harris (1966) has demonstrated that genetic control of enzyme variation in normal people, however, can be rapidly discovered by mass screening methods, and knowledge of inherited enzyme diversity in apparently normal people may soon supersede that in the diseased. An additional approach to discovery of inherited enzyme variation has been pharmacogenetics. Like disease, an unusual pharmacological response selects individuals for study who are more likely to have a genetic variant when an enzyme is known to metabolize the drug in question. Conscious pursuit of enzyme diversity may, in time, make it possible to describe an “enzyme profile” of an individual as distinctive as his finger prints or blood group. The genetics of esterases in man to be described has been studied by both the “mass screening” and the “unusual pharmacological response” methods.

Localization and linkage of three polymorphic DNA sequences on human chromosome 20

The D20S6 locus has been sublocalized by in situ hybridization using the pD3H12 probe to human chromosome band 20p12 and the D20S4 locus using the pMS1-27 probe to 20q13.2. A rare new restriction fragment length polymorphism detected in MspI-digested DNA by the pMSI-27 probe is reported. Linkage studies in nine families have shown that the D20S6 locus is linked to D20S5 (formerly mapped to 20p12 by in situ hybridization) with a maximum likelihood estimate of 0.07 for the recombination frequency (lod score = 9.07) and a confidence interval of 0.02 to 0.14. Estimated recombination frequencies were similar in males and females. Using both two- and multipoint analyses, linkage of D20S4 with the D20S5 and D20S6 loci was excluded and the suggested order for the three loci on chromosome 20 is D20S5-D20S6-centromere-D20S4. D20S5 and D20S6 are very useful markers for linkage studies because of their close proximity and reasonably good polymorphic information content values.