Brigitte F. Brandriff

Chromosomes of human sperm: variability among normal individuals.

SummaryThe chromosomal constitution of 2468 human sperm cells been investigated by fusion of human sperm with hamster eggs. The overall frequency of cells with structural aberrations was 7.7%, ranging from 1.9% to 15.8%, and varying significantly among individuals. The highest frequency occurred in sperm from the oldest donor (49 years), who also had had a vasectomy reversal three years prior to sampling. The overall aneuploidy frequency was 1.7%, ranging from 0.6% to 3.1%. In nine out of ten donors from whom blood samples were available the frequency of sperm cells with structural aberrations was higher than that for lymphocytes. Two previously reported donors (Brandriff et al. 1984) were resampled after an interval of 14 and 16 months respectively, and were each found to have similar frequencies of sperm chromosome abnormalities at both sampling times. A father-son pair included in the study had several chromosome breakpoints in common, although no more frequently than unrelated individuals.

Chromosomal abnormalities in human sperm: comparisons among four healthy men.

SummaryWe have used the human-sperm/hamster-egg system to compare the frequencies of structural and numerical chromosomal aberrations in 909 sperm karyotypes from four normal healthy men. The frequency of structural aberrations was 1.3, 4.8, 9.0, and 10.4% respectively in the four donors. Certain specific breakpoints were seen twice or even three times in three of the donors. The incidence of aneuploidy was 1.3, 1.4, 1.4, and 1.9%. In three donors the frequencies of structural aberrations were significantly higher in sperm than in lymphocytes from the same man. X-to-Y ratios did not differ significantly from the expected 50:50.

Genomic Organization, Chromosomal Localization, Tissue Distribution, and Biophysical Characterization of a Novel MammalianShaker-related Voltage-gated Potassium Channel, Kv1.7

We report the isolation of a novel mouse voltage-gated Shaker-related K+ channel gene,Kv1.7 (Kcna7/KCNA7). Unlike other known Kv1 family genes that have intronless coding regions, the protein-coding region of Kv1.7 is interrupted by a 1.9-kilobase pair intron. The Kv1.7 gene and the related Kv3.3(Kcnc3/KCNC3) gene map to mouse chromosome 7 and human chromosome 19q13.3, a region that has been suggested to contain a diabetic susceptibility locus. The mouse Kv1.7 channel is voltage-dependent and rapidly inactivating, exhibits cumulative inactivation, and has a single channel conductance of 21 pS. It is potently blocked by noxiustoxin and stichodactylatoxin, and is insensitive to tetraethylammonium, kaliotoxin, and charybdotoxin. Northern blot analysis reveals ∼3-kilobase pair Kv1.7transcripts in mouse heart and skeletal muscle. In situhybridization demonstrates the presence of Kv1.7 in mouse pancreatic islet cells. Kv1.7 was also isolated from mouse brain and hamster insulinoma cells by polymerase chain reaction.

Chromosomal damage in sperm of patients surviving Hodgkin's disease following MOPP (nitrogen mustard, vincristine, procarbazine, and prednisone) therapy with and without radiotherapy

Following fusion with hamster eggs, human sperm chromosomes from six Hodgkins disease patients were analyzed to determine the genotoxic effects of therapy. Each patient had received two to six cycles of MOPP (nitrogen mustard, vincristine, procarbazine, and prednisone), with or without radiotherapy, from 3 to 20 years before the study. A total of 571 cells from the six patients were analyzed; 9.8% of the cells had structural aberrations, and 1.6% were hyperhaploid. Analysis of 5998 metaphases from a control group of 24 male donors revealed only 6.9% of cells with structural aberrations and 0.8% aneuploidy. The increase in hyperhaploidy in the patients was statistically significant. Thus, results of this study suggest that the MOPP regimen, with or without radiotherapy, is capable of causing chromosome abnormalities in the sperm of Hodgkins disease patients.

An analysis of structural aberrations in human sperm chromosomes

We have analyzed structural aberrations in 5,000 sperm chromosome complements obtained from 20 men over a 5-yr period by fusion of human sperm with hamster eggs. Detailed data are presented on 366 abnormal cells with 379 analyzable breakpoints. The frequency of cells with structural aberrations ranged from 1.9% to 14.5% among donors; this interindividual variability was statistically significant (p less than 0.0001). In contrast, repeat samples from individual men showed no significant variation over time. The number of sperm chromosome sets processed per hamster egg had no effect on the frequency with which structural aberrations occurred, nor were sperm chromosome abnormalities altered by varying capacitation or culture conditions. The spectrum of structural aberrations observed in human sperm chromosomes and a chi-square analysis of breakpoints based on DNA content are presented. Although human sperm chromosome abnormalities were visualized with a cross-species system, we believe that they represent an inherent, biologically significant phenomenon.

A new system for high-resolution DNA sequence mapping in interphase pronuclei ☆

Cosmid clones containing human or hamster inserts have been hybridized in situ and localized with fluorescent reporter molecules in interphase nuclei (pronuclei) obtained after fusion of hamster eggs with either human or hamster sperm. Hamster egg cytoplasm processes the tightly packaged sperm DNA into large diffuse networks of chromatin fiber bundles, providing hybridization targets more extended than those available in somatic interphase cell nuclei. Pronuclear physical distances between hybridization signals were measured in micrometers and correlated to genomic distances determined by restriction fragment analyses, using cosmids from the Chinese hamster DHFR region and from the human Factor VIII/color vision pigment gene region (Xq28). The mean pronuclear distances between hybridization sites were about three times as large as those measured in somatic interphase cells for equivalent genomic distances. The relationship between physical and genomic distances was linear from less than 50 kb to at least 800 kb. The results show that physical distance in the sperm-egg system promises to extend the mapping range obtainable in somatic interphase nuclei below 50 kb and up to at least 800 kb.

Order and genomic distances among members of the carcinoembryonic antigen (CEA) gene family determined by fluorescence in situ hybridization

Fluorescence in situ hybridization was used to establish the order of, and to estimate genomic distances among, members of the carcinoembryonic antigen (CEA) and pregnancy-specific glycoprotein (PSG) subgroups on chromosome 19. Fluorescence in situ hybridization to metaphase chromosomes localized the PSG subgroup telomeric to the CEA subgroup. Cosmid clones containing sequences for individual genes in the CEA and PSG subgroups were also hybridized to human sperm pronuclear and somatic interphase nuclear chromatin targets. The mapping results lead to the gene order cen-CGM7-CEA-NCA-CGM1-BGP-CGM9-CGM8-PSG-te l. The genomic distances between selected pairs of gene family members were estimated from the physical distances between hybridization sites measured in pronuclei. The CEA-PSG gene family region is estimated to span 1.1 to 1.2 Mb.

The fucosyltransferase locus FUT1 maps distal to apolipoprotein loci E and C2 on human chromosome 19

The location of the fucosyltransferase locus FUT1 relative to the apolipoprotein E/C2 loci on human chromosome 19 has remained unclear. We determined by a combination of physical mapping and fluorescence in situ hybridization that this fucosyltransferase gene maps distal to the apolipoprotein loci.

Assignment of the human lens fiber cell MP19 gene (LIM2) to chromosome 19q13.4, and adjacent to ETFB

The LIM2 gene may play a major role in lens fiber cell structure or communication, and thus cataractogenesis. A human cDNA encoding the corresponding lens fiber cell intrinsic protein MP19 has been previously isolated and characterized. This cDNA had been mapped to human chromosome 19. We have independently confirmed this assignment and fine mapped it to 19q13.4. The position of the LIM2 gene appears to be within 40 kb of the electron transport flavoprotein gene (ETFB) as a cosmid containing sequences from both genes has been identified.

The chromosomal constitution of human sperm selected for motility**Performed under the auspices of the Office of Health and Environmental Research, United States Department of Energy, by the Lawrence Livermore National Laboratory under contract number W-7405-ENG-48.

The chromosome constitutions of sperm selected for motility according to the swim-up technique were compared cytogenetically with those of sperm remaining in the semen with the use of the human sperm/hamster egg system, in which human sperm are fused with hamster eggs to give analyzable haploid chromosome complements. Three semen samples from one donor resulted in 153 chromosome complements from selected, highly motile sperm and 110 unselected, control complements. Four samples were donated by another man, from which 181 selected and 186 control complements were obtained. The frequencies of chromosomal aberrations recovered from the population selected for high motility and the unselected population were not statistically different from one another.