Stochasticity in genetics and gene regulation

Development from fertilized egg to functioning multi-cellular organism requires precision. There is no precision, and often no survival, without plasticity. Plasticity is conferred partly by stochastic variation, present inherently in all biological systems. Gene expression levels fluctuate ubiquitously through transcription, alternative splicing, translation and turnover. Small differences in gene expression are exploited to trigger early differentiation, conferring distinct function on selected individual cells and setting in motion regulatory interactions. Non-selected cells then acquire new functions along the spatio-temporal developmental trajectory. The differentiation process has many stochastic components. Meiotic segregation, mitochondrial partitioning, X-inactivation and the dynamic DNA binding of transcription factor assemblies - all exhibit randomness. Non-random X-inactivation generally signals deleterious X-linked mutations. Correct neural wiring, such as retina to brain, arises through repeated confirmatory activity of connections made randomly. In immune system development, both B-cell antibody generation and the emergence of balanced T-cell categories begin through stochastic trial and error followed by functional selection. Aberrant selection processes lead to immune dysfunction. DNA sequence variants also arise through stochastic events: some involving environmental fluctuation (radiation or presence of pollutants), or genetic repair system malfunction. The phenotypic outcome of mutations is also fluid. Mutations may be advantageous in some circumstances, deleterious in others. This article is part of a discussion meeting issue 'Causes and consequences of stochastic processes in development and disease'

PAX6, brain structure and function in human adults: Advanced MRI in aniridia

Objective PAX6 is a pleiotropic transcription factor essential for the development of several tissues including the eyes, central nervous system, and some endocrine glands. Recently it has also been shown to be important for the maintenance and functioning of corneal and pancreatic tissues in adults. We hypothesized that PAX6 is important for the maintenance of brain integrity in humans, and that adult heterozygotes may have abnormalities of cortical patterning analogous to those found in mouse models. Methods We used advanced magnetic resonance imaging techniques, including surface-based morphometry and region-of-interest analysis in adult humans heterozygously mutated for PAX6 mutations (n = 19 subjects and n = 21 controls). Using immunohistochemistry, we also studied PAX6 expression in the adult brain tissue of healthy subjects (n = 4) and patients with epilepsy (n = 42), some of whom had focal injuries due to intracranial electrode track placement (n = 17). Results There were significant reductions in frontoparietal cortical area after correcting for age and intracranial volume. A greater decline in thickness of the frontoparietal cortex with age, in subjects with PAX6 mutations compared to controls, correlated with age-corrected, accelerated decline in working memory. These results also demonstrate genotypic effects: those subjects with the most severe genotypes have the most widespread differences compared with controls. We also demonstrated significant increases in PAX6-expressing cells in response to acute injury in the adult human brain. Interpretation These findings suggest a role for PAX6 in the maintenance and consequent functioning of the adult brain, homologous to that found in other tissues. This has significant implications for the understanding and treatment of neurodegenerative diseases

Identification of Novel Craniofacial Regulatory Domains Located far Upstream of SOX9 and Disrupted in Pierre Robin Sequence.

Mutations in the coding sequence of SOX9 cause campomelic dysplasia (CD), a disorder of skeletal development associated with 46,XY disorders of sex development (DSDs). Translocations, deletions, and duplications within a ∼2 Mb region upstream of SOX9 can recapitulate the CD-DSD phenotype fully or partially, suggesting the existence of an unusually large cis-regulatory control region. Pierre Robin sequence (PRS) is a craniofacial disorder that is frequently an endophenotype of CD and a locus for isolated PRS at ∼1.2-1.5 Mb upstream of SOX9 has been previously reported. The craniofacial regulatory potential within this locus, and within the greater genomic domain surrounding SOX9, remains poorly defined. We report two novel deletions upstream of SOX9 in families with PRS, allowing refinement of the regions harboring candidate craniofacial regulatory elements. In parallel, ChIP-Seq for p300 binding sites in mouse craniofacial tissue led to the identification of several novel craniofacial enhancers at the SOX9 locus, which were validated in transgenic reporter mice and zebrafish. Notably, some of the functionally validated elements fall within the PRS deletions. These studies suggest that multiple noncoding elements contribute to the craniofacial regulation of SOX9 expression, and that their disruption results in PRS

Monoallelic variants resulting in substitutions of MAB21L1 Arg51 Cause Aniridia and microphthalmia

Classical aniridia is a congenital and progressive panocular disorder almost exclusively caused by heterozygous loss-of-function variants at the PAX6 locus. We report nine individuals from five families with severe aniridia and/or microphthalmia (with no detectable PAX6 mutation) with ultrarare monoallelic missense variants altering the Arg51 codon of MAB21L1. These mutations occurred de novo in 3/5 families, with the remaining families being compatible with autosomal dominant inheritance. Mice engineered to carry the p. Arg51Leu change showed a highly-penetrant optic disc anomaly in heterozygous animals with severe microphthalmia in homozygotes. Substitutions of the same codon (Arg51) in MAB21L2, a close homolog of MAB21L1, cause severe ocular and skeletal malformations in humans and mice. The predicted nucleotidyltransferase function of MAB21L1 could not be demonstrated using purified protein with a variety of nucleotide substrates and oligonucleotide activators. Induced expression of GFP-tagged wildtype and mutant MAB21L1 in human cells caused only modest transcriptional changes. Mass spectrometry of immunoprecipitated protein revealed that both mutant and wildtype MAB21L1 associate with transcription factors that are known regulators of PAX6 (MEIS1, MEIS2 and PBX1) and with poly(A) RNA binding proteins. Arg51 substitutions reduce the association of wild-type MAB21L1 with TBL1XR1, a component of the NCoR complex. We found limited evidence for mutation-specific interactions with MSI2/Musashi-2, an RNA-binding proteins with effects on many different developmental pathways. Given that biallelic loss-of-function variants in MAB21L1 result in a milder eye phenotype we suggest that Arg51-altering monoallelic variants most plausibly perturb eye development via a gain-of-function mechanism

Stepped Care for Maternal Mental Health: A Case Study of the Perinatal Mental Health Project in South Africa

As one article in a series on Global Mental Health Practice, Simone Honikman and colleagues from South Africa provide a case study of the Perinatal Mental Health Project, which delivered mental health care to pregnant women in a collaborative, step-wise manner, making use of existing resources in primary care

Linkage of the structural gene for uroporphyrinogen I synthase to markers on mouse chromosome 9 in a cross between feral and inbred mice

The Ups locus has been mapped to mouse chromosome 9 in a three-point cross. The observed gene order is centromere- Ups -15- Mpi-1 -22- Mod-1 . Ups is unlinked to Lv , which encodes the previous enzyme in the heme biosynthesis pathway. Feral mice collected at Skive, Denmark, have been characterized at several biochemical loci; multiple differences from inbred strains make this a useful stock for linkage analysis.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/44143/1/10528_2004_Article_BF00483967.pd

Renal malformations associated with mutations of developmental genes: messages from the clinic

Renal tract malformations (RTMs) account for about 40% of children with end-stage renal failure. RTMs can be caused by mutations of genes normally active in the developing kidney and lower renal tract. Moreover, some RTMs occur in the context of multi-organ malformation syndromes. For these reasons, and because genetic testing is becoming more widely available, pediatric nephrologists should work closely with clinical geneticists to make genetic diagnoses in children with RTMs, followed by appropriate family counseling. Here we highlight families with renal cysts and diabetes, renal coloboma and Fraser syndromes, and a child with microdeletion of chromosome 19q who had a rare combination of malformations. Such diagnoses provide families with often long-sought answers to the question “why was our child born with kidney disease”. Precise genetic diagnoses will also help to define cohorts of children with RTMs for long-term clinical outcome studies

CTCF cis-Regulates Trinucleotide Repeat Instability in an Epigenetic Manner: A Novel Basis for Mutational Hot Spot Determination

At least 25 inherited disorders in humans result from microsatellite repeat expansion. Dramatic variation in repeat instability occurs at different disease loci and between different tissues; however, cis-elements and trans-factors regulating the instability process remain undefined. Genomic fragments from the human spinocerebellar ataxia type 7 (SCA7) locus, containing a highly unstable CAG tract, were previously introduced into mice to localize cis-acting “instability elements,” and revealed that genomic context is required for repeat instability. The critical instability-inducing region contained binding sites for CTCF—a regulatory factor implicated in genomic imprinting, chromatin remodeling, and DNA conformation change. To evaluate the role of CTCF in repeat instability, we derived transgenic mice carrying SCA7 genomic fragments with CTCF binding-site mutations. We found that CTCF binding-site mutation promotes triplet repeat instability both in the germ line and in somatic tissues, and that CpG methylation of CTCF binding sites can further destabilize triplet repeat expansions. As CTCF binding sites are associated with a number of highly unstable repeat loci, our findings suggest a novel basis for demarcation and regulation of mutational hot spots and implicate CTCF in the modulation of genetic repeat instability