Download Genetics Examination question

Some questions for discussion paper 3
Note: You do not need to prepare these for turn in, but they will be used as a guide for the
class discussion. Other questions, comments and points of discussion are also most welcome!
Edery P, Marcaillou C, Sahbatou M, Labalme A, Chastang J, Touraine R, Tubacher E, Senni F,
Bober MB, Nampoothiri S, Jouk PS, Steichen E, Berland S, Toutain A, Wise CA, Sanlaville D,
Rousseau F, Clerget-Darpoux F, Leutenegger AL (2011) Science 332:240-3.
Association of TALS developmental disorder with defect in minor splicing component U4atac
snRNA.
Pessa, HKJ and MK Frilander (2011) Minor splicing, disrupted. Science 332:184-185
1- List any terms or techniques used in this paper that are unfamiliar to you, and do some quick
web searching to become more familiar with them.
Homozygosity mapping
Autozygous
2 This paper utilized the technique of homozygosity mapping to map the mutation that
conditions microcephalic osteodysplastic primordial dwarfism type I (MOPD I), a severe
developmental disorder, also known as Taybi-Linder syndrome (TALS). How does homozygosity
mapping work?
Markers linked to a recessive genetic mutation are identified by the fact that these markers are
homozygous [5 pt] and identical by descent (IBD) [5 pt] from both parents in affected, inbred
individuals. The strategy is to identify haplotypes that are homozygous and IBD in affected
individuals and heterozygous in unaffected relatives [5 pt]
3 Describe the types of mutations and families that are suited to homozygosity mapping and
explain how or why these types of mutations and families are essential to the success of this
mapping approach.
1-The mutations must be recessive [5 pt] so that affected individuals are expected to be
homozygous for the mutation and linked marker alleles [5 pt].
2-The mutation must also be rare in the population as a whole [5 pt], and
3-The segregating families must have some degree of inbreeding 5 pt], so that the probability
of homozygosity for the mutation through IBD is much greater than the probability of
homozygosity for the mutation by random chance [5 pt]. (this point missed by most)
4 Once the TALS mutation was localized to chromosome 2q14.2, this group encountered
challenges in their attempts to identify the TALS gene and causal mutations. Explain how this
goal was initially approached, why this approach failed, and how the TALS gene and mutations
were ultimately identified. What criteria were used for identification of the causal mutations.
Both groups initially Sanger sequenced (from selected individuals) exons of annotated, proteincoding genes in the 2q14 region [2.5 pt]. This failed to reveal likely disease-causing mutations
because the TALS locus is not protein-coding [2.5 pt]! Both groups turned to high-throughput
sequencing of all non-repetitive DNA within the region associated with TALS through
homozygosity mapping 2.5 pt]. The criteria for identification of the causal mutation were: 1homozygosity in affected individuals [2.5 pt]; 2-heterozygosity in parents of affected individuals
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Some questions for discussion paper 3
Note: You do not need to prepare these for turn in, but they will be used as a guide for the
class discussion. Other questions, comments and points of discussion are also most welcome!
[2.5 pt] 3-absent from healthy sibs that did not carry the 2q14.2 haplotype homozygous in
TALS-affected individuals [2.5 pt] 4-rare or previously unknown SNP [2.5 pt].
5 What is the nature of the TALS gene? Where was it located with respect to neighboring
genes? What are the implications of this finding for others seeking to identify genes through
map and genome-based approaches?
In both studies, the only DNA sequence differences meeting the homozygosity mapping criteria
fell within the U4atac small nuclear RNA (snRNA) gene that encodes and RNA component of the
minor, U12, spliceosome [2.5 pt]. Therefore, searches for causal mutations should not be
limited to protein coding genes [2.5 pt]. (Unfortunately protein-coding sequences are the first,
and generally best annotated regions of sequenced genomes.)
6 What are the types of mutations that were identified as causal to TALS? Not every individual
affected with TALS was homozygous for a given TALS mutation. Explain why these individuals
were still affected.
The TALS mutations were single base changes [2.5 pt] that appeared to alter 5' or 3' stem loops
essential to spliceosome function [2.5 pt].
He et al. also looked at two German families in which affected individuals had very similar
phenotypes to those seen in the Ohio Amish. In the German families, however, affected
individuals were compound heterozygotes [5 pt]: 55G>A 111G / 55G 111G>A. Since the
two mutations fall into the same unit of function, they are non-complementing[5 pt] and a
mutant phenotype results.
7 Considering the nature of the TALS gene and mutations, discuss the probable molecular and
cellular basis for TALS and the data that support such a basis. What questions remain
unresolved with respect to the molecular and cellular basis of this disease? Suggest an
experimental approach that might be used to address these questions.
TALS is likely to result from failed activity of the minor splicosome, so even though the U12
pathway does not splice a lot of introns, correct splicing of at least some U12-dependent
introns is clearly essential to normal growth and development [2.5 pt]. Fibroblast cell lines
homozygous for 51G>A show low ratios of spliced to un-spliced U12-dependent introns
compared to U2 (major splicosome) dependent introns;
8) Although there is still lots to be learned about TALS, what are some of the immediate
practical applications of this study? Genetic testing for carriers [2.5 pt], correct clinical diagnosis
of developmental disorders [2.5 pt], potential for the development of gene therapy [2.5 pt].
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