Unlinked Families Research Articles

Linkage mapping of the spinal muscular atrophy gene

Spinal muscular atrophy (SMA) is a common autosomal recessive disorder resulting in loss of motor neurons. We have performed linkage analysis on a panel of families using nine markers that are closely linked to the SMA gene. The highest lod score was obtained with the marker D5S351 (Zmax = 10.04 at theta = O excluding two unlinked families, and Zmax = 8.77 at theta = 0.007 with all families). One type III family did not show linkage to the 5q13 markers, and in one type I consanguineous family the affected individual did not show homozygosity except for the marker D5S435. Three recombinants were identified with the closet centromeric marker, D5S435, which position the gene telomeric of this marker. These recombinants will facilitate finer mapping of the location of the SMA gene. Lastly, two families provide strong evidence for a remarkable variability in presentation of the SMA phenotype, with the age at onset in one family varying from 17 months to 13 years.

Genetic heterogeneity of Usher syndrome type II.

Usher syndrome is an autosomal recessive disorder characterised by retinitis pigmentosa and congenital sensorineural hearing loss. A gene for Usher syndrome type II (USH2) has been localised to chromosome 1q32-q41. DNA from a family with four of seven sibs affected with clinical characteristics of Usher syndrome type II was genotyped using markers spanning the 1q32-1q41 region. These included D1S70 and D1S81, which are believed to flank USH2. Genotypic results and subsequent linkage analysis indicated non-linkage of this family to these markers. The A test analysis for heterogeneity with this family and 32 other Usher type II families was statistically significant at p < 0.05. Further clinical evaluation of this family was done in light of the linkage results to determine if any phenotypic characteristics would allow for clinical identification of the unlinked type. No clear phenotypic differences were observed; however, this unlinked family may represent a previously unreported subtype of Usher type II characterised by a milder form of retinitis pigmentosa and mild vestibular abnormalities. Heterogeneity of Usher syndrome type II complicates efforts to isolate and clone Usher syndrome genes using linkage analysis and limits the use of DNA markers in early detection of Usher type II.

Autosomal dominant polycystic kidney disease with minimal clinical expression unlinked to the PKD1 locus.

Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutations at the PKD1 locus in most families. This locus has been assigned to the short arm of chromosome 16 by linkage analysis. It has been estimated that approximately 5% of families have a disease that does not map to this locus and most of these families have clinical features indistinguishable from the disease caused by PKD1 mutations. We report a large three-generation Caucasian family from Northern Ireland with ADPKD in whom all affected individuals (age range 22-68) were normotensive and only the two eldest had mild renal impairment. Linkage was excluded between the disease and both the alpha-globin gene complex and the microsatellite marker D16S283. This family confirms that phenotypic heterogeneity exists between unlinked families and that certain non-PKD1 mutations cause mild disease expression. Many such individuals may therefore remain undetected and the incidence of families with ADPKD who have non-PKD1 mutations may be greater than previously estimated.

A bootstrap approach to estimating power for linkage heterogeneity.

We examined the power of detecting linkage heterogeneity when the null hypothesis is that all families are linked to one locus (A) and the two alternative hypotheses are either 1) a proportion of the families are linked to locus A and the remaining families are linked to a second locus B or 2) a proportion of the families are linked to locus A or B and a third proportion of the families are unlinked to either locus. The power of detecting linkage heterogeneity is estimated for various proportions of families linked to loci A, B or unlinked to either locus (sampling under the alternative hypothesis). To estimate the significance level, the data set is sampled under the null hypothesis. For sampling under both hypotheses, a bootstrap approach is employed, sampling the simulated pedigrees with replacement. The power to detect linkage heterogeneity is strongest when the recombination fraction is 0 and equal proportions of the families are linked to loci A and B. The power decreases as the recombination fraction increases, the proportion of unlinked families increases and the disparity between the proportion of the families linked to either locus A or B increases. In the data set of 32 Duke Familial Alzheimer Disease families, when equal proportions of families are linked to loci A and B, the power to detect linkage heterogeneity is 0.94 using a likelihood ratio criterion of 10:1. The p value that corresponds to the likelihood ratio of 10:1 is estimated as 0.013 with a 95% confidence interval for p ranging from 0.012 to 0.014.

Multipoint mapping of adult onset polycystic kidney disease (PKD1) on chromosome 16.

Analysis of genetic linkage data in 33 adult onset polycystic kidney (ADPKD) families was carried out using probes for the D16S85, D16S84, and D16S94 loci. The data set of 33 families shows no evidence of genetic heterogeneity since one unlinked family was previously excluded. Two point linkage analysis showed maximum likelihood values of the recombination fraction of 0.07 for ADPKD and D16S85 (lod score 18.78), 0.02 for ADPKD and D16S84 (lod score 7.55), and 0.00 for ADPKD and D16S94 (lod score 6.73). Multipoint analysis showed a maximum likelihood order of tel-D16S85-0.06-D16S84-0.02-(PKD1, D16S94)-cen with a multipoint lod score of 32.16. Analysis of rare recombinants lying close to PKD1 gave results consistent with this order.

Evidence for genetic heterogeneity in hereditary hydronephrosis caused by pelvi-ureteric junction obstruction, with one locus assigned to chromosome 6p.

Hereditary hydronephrosis (MIM 143400) is an autosomal dominant trait that causes unilateral or bilateral pelvi-ureteric junction (PUJ) obstruction. Linkage analysis was undertaken in 5 families with hereditary PUJ obstruction using the major histocompatibility complex locus as a test marker. The data as a whole supported a hereditary hydronephrosis locus on 6p. Maximal lod scores were 3.090 at a recombination fraction of 0.1 with full penetrance, and 2.486 at a recombination fraction of 0.1 with a penetrance of 90%. However, analysis of two point lod scores using the HOMOG program revealed significant evidence for genetic heterogeneity with one locus on 6p in 4 of the families, and a different locus in one family. After exclusion of this unlinked family, two point analysis gave a maximal lod score of 3.9 at a recombination fraction of 0.05 with full penetrance, and 4.2 at a recombination fraction of 0.0 with 90% penetrance. These data support the assignment of one of the loci for hereditary hydronephrosis to chromosome 6p.

Effect of heterogeneity and assumed mode of inheritance on lod scores.

Heterogeneity is a major factor in many common, complex diseases and can confound linkage analysis. Using computer-simulated heterogeneous data we tested what effect unlinked families have on a linkage analysis when heterogeneity is not taken into account. We created 60 data sets of 40 nuclear families each with different proportions of linked and unlinked families and with different modes of inheritance. The ascertainment probability was 0.05, the disease had a penetrance of 0.6, and the recombination fraction for the linked families was zero. For the analysis we used a variety of assumed modes of inheritance and penetrances. Under these conditions we looked at the effect of the unlinked families on the lod score, the evaluation of the mode of inheritance, and the estimate of penetrance and of the recombination fraction in the linked families. 1. When the analysis was done under the correct mode of inheritance for the linked families, we found that the mode of inheritance of the unlinked families had minimal influence on the highest maximum lod score (MMLS) (i.e., we maximized the maximum lod score with respect to penetrance). Adding sporadic families decreased the MMLS less than adding recessive or dominant unlinked families. 2. The mixtures of dominant linked families with unlinked families always led to a higher MMLS when analyzed under the correct (dominant) mode of inheritance than when analyzed under the incorrect mode of inheritance. In the mixtures with recessive linked families, assuming the correct mode of inheritance generally led to a higher MMLS, but we observed broad variation.(ABSTRACT TRUNCATED AT 250 WORDS)

Recent Linkage Studies in Tuberous Sclerosis

After our initial reports 3-5 supporting a locus for tuberous sclerosis (TSC) on 9q, linkage analysis was undertaken in eight large multigeneration TSC families using nine polymorphic markers. Six of the markers were from the distal long arm of chromosome 9 and three from the long arm of chromosome 11. The data as a whole supported a TSC locus on distal 9q, the peak lod score on multipoint analysis being 3.77 6 cM proximal to the Abelson oncogene locus (ABL). However, analysis of 2-point lod scores using the HOMOG programs revealed significant evidence for genetic heterogeneity (p = 0.01), tight linkage to ABL being highly unlikely in one family. After exclusion of the unlinked family multipoint, analysis gave a peak lod score of 6.1 in the vicinity of ABL. The family unlinked to ABL showed no recombinants with two chromosome 11 probes, but it was too small to provide significant evidence for linkage. Genetic heterogeneity in TSC as demonstrated by this study will complicate efforts to clone the genes by reverse genetics and will severely hamper the use of linked probes for carrier detection and prenatal diagnosis.

Phenotypic heterogeneity of spinal muscular atrophy mapping to chromosome 5q11.2‐13.3 (SMA 5q)

We made phenotypic analysis of 14 families with spinal muscular atrophy (SMA) linking to chromosome 5q11.2-13.3 (SMA 5q), and 2 that may not map to this locus, to assess clinical symptoms among SMA families known to result from mutation at the identical gene/locus. Although the current number of families is still small, the correlation of clinical phenotype and molecular genotype supports 2 observations. First, SMA mutations at the 5q locus present with a broad continuum of clinical abnormalities, and 2nd, the single clearly unlinked family presents with an unusual phenotype characterized by relatively late onset and early death. Thus, there are as yet no unambiguous cases of typical SMA families that are clearly unlinked to the locus at 5q-ie, no clear cases of nonallelic heterogeneity. Analysis of SMA 5q families supports the view that, with certain exceptions, there is little phenotypic intrafamilial variability. When families were ranked by severity of disease there was a strong correlation with age of onset. Onset within the 1st few months was associated with early death, but not in all cases. With rare exception, onset after 1 year of age was associated with less severe disease and greater longevity.

Evidence for genetic heterogeneity in tuberous sclerosis.

The question of genetic heterogeneity in tuberous sclerosis (TSC) was addressed by genetic linkage studies in eight affected families using nine polymorphic markers (EFD126.3, MCT136, ABO, ABL, AK1, and MCOA12 from distal 9q, and PBGD, MCT128.1, and 1CJ52.208M from distal 11q). The data as a whole supported a TSC locus on distal 9q, the peak lod score on multipoint analysis being 3.77 at 6 cM proximal to the Abelson oncogene locus (ABL). However, analysis of two point lod scores using the HOMOG programs showed significant evidence for genetic heterogeneity (p = 0.01), linkage to ABL being unlikely in one family. After exclusion of the unlinked family, multipoint analysis gave a peak lod score of 6.1 in the vicinity of ABL. The family unlinked to ABL showed no recombinants with two chromosome 11 probes, but was too small to provide significant evidence for linkage. Genetic heterogeneity in TSC will complicate efforts to clone the causative genes and severely limit the use of linked probes for carrier detection and prenatal diagnosis.

Genetic prediction in X-linked agammaglobulinaemia.

S21 (DXS17) and pXG12 (DXS94), two probes linked to the locus of X-linked agammaglobulinaemia (XLA), were used for genetic prediction in 13 such families. A method of allowing for nonallelic genetic heterogeneity was demonstrated in the calculation of the genetic risks, specifying a certain proportion of unlinked families. We further estimated the impact due to the uncertainty of the proportion of unlinked families on the final genetic risks in each family and this can be taken into account during genetic counselling.

Rho-dependent transcription termination of a bacterial operon is antagonized by an extrachromosomal gene product.

The psu gene product of "phasmid" (phage-plasmid) P4 acts as a transcription antitermination factor in trans and in cis, respectively, within the morphogenic operons of its P2 phage helper during lytic viral development and on P4 itself during the establishment stage of its alternative mode of propagation as a plasmid. Here we show that psu also antagonizes activity of the Escherichia coli transcription termination factor rho at the terminator of the trp operon. Such a finding provides to our knowledge the first direct evidence for antitermination activity at a known rho-dependent site by the psu gene product. It also reveals an example of an extrachromosomal gene product that acts on specific sites of three different genomes to regulate expression of unlinked families of genes.

Multiple alpha and beta tubulin genes represent unlinked and dispersed gene families.

Cloned cDNA sequences specific for alpha or beta tubulin mRNAs have been used to show that the multigene families which encode either alpha or beta tubulin are unlinked and dispersed throughout the chicken genome. Fractions of chicken chromosomes partially purified by centrifugation on a sucrose gradient were digested with restriction endonucleases and electrophoresed on agarose gels. The DNA was transferred to nitrocellulose filters and hybridized to labeled probes constructed from cloned cDNA sequences specific for alpha or beta tubulin. We find alpha tubulin sequences on four different chicken chromosomes and beta tubulin sequences on at least two different chromosomes. Moreover, using chicken chromosomes further purified with a fluorescent cell sorter, we have been able unambiguously to localize alpha tubulin genes to chromosome 1 and chromosome 8 and two of the beta genes to chromosome 2.