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Cornelia de Lange syndrome
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Drosophila genes associated with
Cornelia de Lange syndrome
Nipped-B
Related terms

Mitosis
Overview of the disease

Cornelia de Lange Syndrome (CdLS) alters many aspects of growth and development. CdLS is caused by mutations in genes encoding proteins that ensure that chromosomes are distributed equally when a cell divides. These include genes that encode components of the cohesin complex, and Nipped-B-Like (NIPBL) that puts cohesin onto chromosomes. Individuals with CdLS have only modest reductions in the activities of these genes and do not show changes in chromosome distribution. Instead, they show differences in the expression many genes that control development. Drosophila, with one mutant copy of the Nipped-B gene, which is equivalent to the NIPBL gene, show characteristics similar to individuals with CdLS (Wu, 2015 and references therein).

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Relevant studies of Cornelia de Lange syndrome

Wu, Y., Gause, M., Xu, D., Misulovin, Z., Schaaf, C.A., Mosarla, R.C., Mannino, E., Shannon, M., Jones, E., Shi, M., Chen, W.F., Katz, O.L., Sehgal, A., Jongens, T.A., Krantz, I.D. and Dorsett, D. (2015). Drosophila Nipped-B mutants model Cornelia de Lange syndrome in growth and behavior. PLoS Genet 11: e1005655. PubMed ID: 26544867

Abstract
Individuals with Cornelia de Lange Syndrome (CdLS) display diverse developmental deficits, including slow growth, multiple limb and organ abnormalities, and intellectual disabilities. Severely-affected individuals most often have dominant loss-of-function mutations in the Nipped-B-Like (NIPBL) gene, and milder cases often have missense or in-frame deletion mutations in genes encoding subunits of the cohesin complex. Cohesin mediates sister chromatid cohesion to facilitate accurate chromosome segregation, and NIPBL is required for cohesin to bind to chromosomes. Individuals with CdLS, however, do not display overt cohesion or segregation defects. Rather, studies in human cells and model organisms indicate that modest decreases in NIPBL and cohesin activity alter the transcription of many genes that regulate growth and development. Sister chromatid cohesion factors, including the Nipped-B ortholog of NIPBL, are also critical for gene expression and development in Drosophila melanogaster. This study describes how a modest reduction in Nipped-B activity alters growth and neurological function in Drosophila. These studies reveal that Nipped-B heterozygous mutant Drosophila show reduced growth, learning, and memory, and altered circadian rhythms. Importantly, the growth deficits are not caused by changes in systemic growth controls, but reductions in cell number and size attributable in part to reduced expression of myc (diminutive) and other growth control genes. The learning, memory and circadian deficits are accompanied by morphological abnormalities in brain structure. These studies confirm that Drosophila Nipped-B mutants provide a useful model for understanding CdLS, and provide new insights into the origins of birth defects (Wu, 2015).

Highlights

  • Heterozygous Nipped-B mutations reduce growth to a similar extent as mutations in the dm (myc) and Tor growth regulators.
  • Systemic growth control and developmental timing are not detectably altered in heterozygous Nipped-B mutants.
  • Nipped-B mutant wings have fewer and smaller cells.
  • Ectopic dm expression restores wing size and increases the weight of Nipped-B mutants.
  • Heterozygous Nipped-B mutant wing discs show broad modest decreases in expression of genes supporting development and growth.
  • Heterozygous Nipped-B mutants have abnormal brain structures and show learning and memory deficits.
  • Nipped-B mutants are arrhythmic and sleep less than controls.
  • Heterozygous Nipped-B mutants show no signs of seizures.

Discussion
Prior studies have revealed that Nipped-B heterozygous mutant Drosophila display subtle and latent external morphological phenotypes that become overt in adults only when combined with mutations in key developmental regulatory genes such as cut, Ultrabithorax, Notch, mastermind, hedgehog, and genes encoding cohesin or Polycomb silencing complex subunits. This contrasts with mice and humans, where similar deficiencies in NIPBL cause multiple specific and obvious morphological changes. There are also differences between mice and humans. For instance, limb abnormalities are largely absent in Nipbl(+/-) mice, but heart defects are significantly more frequent than in individuals with CdLS. Extrapolating from Drosophila genetic interaction data, this study posits that the individual physical birth defects in vertebrates stem from altered expression of specific sets of developmental genes, and that the variability between individuals with CdLS reflects differences in genetic background (Wu, 2015).

It was shown that although the external morphological changes in Nipped-B mutant Drosophila are minimal in an otherwise wild-type background, they share the reduced size with Nipbl(+/-) mice and CdLS. The data argue that the reduced size reflects decreases in both total cell number and size, and not changes in the systemic control of growth that sense critical body mass, deficits in the utilization of nutrition, or increased cell death. The decrease in size likely stems in part from modestly reduced expression of the myc (dm), Tor, InR and other genes that promote cell proliferation, division and growth, and not increased apoptosis. Indeed, one of the most intriguing findings is the reduced ability of excess dm expression to induce apoptosis in Nipped-B heterozygous mutants. This may stem in part from reduced function of a Dm-dependent enhancer that drives expression of the grim and reaper pro-apoptosis genes. This region binds Nipped-B and cohesin in wing discs, and deletion of this region permits excess dm expression using the tub-myc driver to increase wing size more than in wild-type flies because it reduced apoptosis (Wu, 2015).

It remains to be seen to what extent these findings in Drosophila might explain the reduced growth in CdLS and in Nipbl(+/-) mice. It seems likely that similar mechanisms at least make a significant contribution to the reduced growth in mammals because Nipbl(+/-) mice and cells from individuals with CdLS also show reduced c-myc expression. Organisms use a variety of mechanisms to sense body size and regulate growth and developmental transitions. Drosophila transitions are timed by pulses of the ecdysone steroid hormone produced by the prothoracic gland located between the two lobes of the brain. Specific neurons in the brain secrete a peptide hormone, prothoracicotropic hormone (PTTH) that stimulates ecdysone production. Insulin signaling, nutrition and many other factors, which are not all well understood, control the hormonal pathways and timing of the ecdysone pulses that determine absolute body size. For example, nutrient deprivation substantially delays the ecdysone pulses, but not enough to fully restore maximal body size. What is particularly striking is how close Nipped-B mutants are to wild type in their developmental timing and in how their developmental staging responds to nutrient deprivation. This argues that the systemic hormonal pathways that regulate body size and hormonal pulses that induce developmental stages are largely unaffected, and that the reduced size of Nipped-B mutants stems primarily from a small but significant reduction in the number of cell divisions, and in the mechanisms that determine final cell size (Wu, 2015).

It is more difficult to precisely time the developmental staging in mice and humans than in Drosophila, and thus to determine whether or not developmental timing or systemic growth pathways are significantly altered with decreased NIPBL function. Although some individuals with CdLS show slightly delayed puberty, puberty occurs at the normal age in many. The slight delays, and incomplete pubertal changes might all be attributed to causes other than a general developmental delay, such as structural abnormalities or changes in hormone levels. The relatively normal timing of puberty, therefore seems to indicates that the more extreme reductions in overall size observed in CdLS are also more likely to stem from changes in cell number and size than changes in systemic body size controls (Wu, 2015).

It was also found that Drosophila Nipped-B heterozygotes display many behavioral and neurological features resembling those seen in CdLS patients: they are deficient in learning and short-term memory, and display disruptive sleep patterns and abnormal circadian rhythms. Intellectual disability is the most common clinical phenotype seen in individuals with CdLS. The average IQ score of typical CdLS cases, mostly with NIPBL mutations, is 53 (range 30–86). It was found that although Nipped-B heterozygous mutant flies are capable of learning, their learning capacity is significantly lower than that of the wiso31 controls. Furthermore, these Nipped-B mutants are accompanied by pleiotropic structural abnormalities in mushroom bodies, the major brain structures controlling learning and memory. While it is conceivable that the morphological defects in the mushroom bodies could contribute to the learning and memory deficits observed in the Nipped-B mutants, how these structural and functional deficiencies correlate with each other warrant further detailed studies (Wu, 2015).

A striking similarity was observed in the sleep patterns of fly Nipped-B heterozygotes and those seen in CdLS patients. Sleep disturbances are common in CdLS and seen in up to 55% of CdLS individuals. Insomnia (difficulty in initiating sleep), difficulty staying asleep, frequent night wakenings, and sleepiness during the daytime are the most common sleep problems reported in CdLS. Frequent but dramatically shortened sleep episodes was the characteristic sleep pattern observed in Nipped-B heterozygote flies. In fact, the reduced length of sleep episodes lead to a dramatic loss in daytime and nighttime sleep, which was unable to be compensated for by significant increases in the number of sleep bouts (Wu, 2015).

While it has been speculated that sleep disturbances in CdLS individuals may be in part attributable to a circadian rhythm disorder, strict and objective studies to confirm this suggestion have not been undertaken. The locomotor activity-based circadian rhythm assay is a well-established measure of circadian rhythm in Drosophila. It was demonstrated that an aberrant circadian rhythm exists in a large fraction of Nipped-B haploinsufficient male and female flies. For Nipped-B mutants that still display rhythmic circadian patterns, their free-running activity rhythms are maintained at around 24 hours, similar to the periodicity of wiso31 controls. A remarkable consistency between the circadian defects and sleep aberrance was observed in these Nipped-B heterozygous mutants; flies that were profoundly arrhythmic were the ones that showed the most disturbed sleep patterns. Nevertheless, the circadian rhythm alterations are more apparent than the sleep disturbances in males, suggesting that additional sex-specific factors are involved in determining the sleep patterns. Taken together, these data suggest that at least in fly Nipped-B mutants, intrinsic circadian rhythm defects likely contribute to their aberrant sleep patterns (Wu, 2015).

Overall, this study on fly Nipped-B mutants demonstrates a strikingly analogous growth and neurocognitive/behavioral phenotype between heterozygous Nipped-B mutants and human CdLS individuals, including small body size, learning and memory deficits, disruptive sleep patterns and circadian rhythm defects. Drosophila Nipped-B heterozygotes are thus a valuable resource, with multiple objective and readily measureable metrics, for modeling human CdLS. Studies on the function of Nipped-B and cohesin components in CdLS patient cell lines, Nipbl(+/-) mouse and zebrafish models, and Drosophila have contributed substantially to our understanding of the roles of Nipped-B and cohesin components in development and gene regulation. The presence of a sophisticated genetic tool kit and economical availability of fruit flies will make it possible to explore developmental deficits in a tissue- and stage-specific manner, as well as to test their relevance to human development and the pathogenesis of CdLS. Drosophila is an ideal model organism to address these issues, given its short life cycle, lower degree of genomic redundancy and the available genetic tools. Nipped-B mutants can also be utilized to search for and test new pharmacologic therapeutic modalities, towards amelioration of the growth and neurodevelopmental functioning of individuals with CdLS (Wu, 2015).

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Krantz, I.D., McCallum, J., DeScipio, C., Kaur, M., Gillis, L.A., Yaeger, D., Jukofsky, L., Wasserman, N., Bottani, A., Morris, C.A., Nowaczyk, M.J., Toriello, H., Bamshad, M.J., Carey, J.C., Rappaport, E., Kawauchi, S., Lander, A.D., Calof, A.L., Li, H.H., Devoto, M. and Jackson, L.G. (2004). Cornelia de Lange syndrome is caused by mutations in NIPBL, the human homolog of Drosophila melanogaster Nipped-B. Nat Genet 36: 631-635. PubMed ID: 15146186

Abstract
Cornelia de Lange syndrome is a dominantly inherited multisystem developmental disorder characterized by growth and cognitive retardation; abnormalities of the upper limbs; gastroesophageal dysfunction; cardiac, ophthalmologic and genitourinary anomalies; hirsutism; and characteristic facial features. Genital anomalies, pyloric stenosis, congenital diaphragmatic hernias, cardiac septal defects, hearing loss and autistic and self-injurious tendencies also frequently occur. Prevalence is estimated to be as high as 1 in 10,000. This study carried out genome-wide linkage exclusion analysis in 12 families with CdLS and identified four candidate regions, of which chromosome 5p13.1 gave the highest multipoint lod score of 2.7. This information, together with the previous identification of a child with CdLS with a de novo t(5;13)(p13.1;q12.1) translocation, allowed delineation of a 1.1-Mb critical region on chromosome 5 for the gene mutated in CdLS. Mutations in one gene in this region were identified, which was named NIPBL, in four sporadic and two familial cases of CdLS. The genomic structure of NIPBL was characterized and it was found that it is widely expressed in fetal and adult tissues. The fly homolog of NIPBL, Nipped-B, facilitates enhancer-promoter communication and regulates Notch signaling and other developmental pathways in Drosophila melanogaster (Krantz, 2004).

Highlights

  • Identification of NIPBL as the gene underlying CdLS.
  • Mutations and clinical features in individuals with CdLS.
  • Expression of NIPBL in the developing mouse.

Discussion
Nipped-B is an essential regulator of cut, Ultrabithorax and Notch receptor signaling. Its protein product belongs to the family of chromosomal adherins, and genetic evidence suggests that it has an architectural role in facilitating long-distance interactions between enhancers and promoters. The involvement of Nipped-B in regulating Notch signaling is of interest, as two other genes involved in Notch signaling are implicated in human developmental disorders (mutations in JAG1 result in Alagille syndrome, and mutations in DLL3 result in spondylocostal dysostosis (Krantz, 2004).

The identification of mutations in a single allele of NIPBL in individuals with CdLS is consistent with a dominant pattern of inheritance. All mutations identified so far predict a truncated protein product and probably result in functional haploinsufficiency. That haploinsufficiency is a mechanism in CdLS is confirmed by the child with a large deletion of the region (encompassing NIPBL) and severe manifestations of CdLS, and by the child with the translocation reported in this study, who also has severe manifestations (Krantz, 2004).

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Tonkin, E.T., Wang, T.J., Lisgo, S., Bamshad, M.J. and Strachan, T. (2004). NIPBL, encoding a homolog of fungal Scc2-type sister chromatid cohesion proteins and fly Nipped-B, is mutated in Cornelia de Lange syndrome. Nat Genet 36: 636-641. PubMed ID: 15146185

Abstract
Cornelia de Lange syndrome (CdLS) is a multiple malformation disorder characterized by dysmorphic facial features, mental retardation, growth delay and limb reduction defects. This study identified and characterized a new gene, NIPBL, that is mutated in individuals with CdLS and determined its structure and the structures of mouse, rat and zebrafish homologs. Its protein product was named delangin. Vertebrate delangins have substantial homology to orthologs in flies, worms, plants and fungi, including Scc2-type sister chromatid cohesion proteins, and D. melanogaster Nipped-B. The study proposes that perturbed delangin function may inappropriately activate DLX genes, thereby contributing to the proximodistal limb patterning defects in CdLS. Genome analyses typically identify individual delangin or Nipped-B-like orthologs in diploid animal and plant genomes. The evolution of an ancestral sister chromatid cohesion protein to acquire an additional role in developmental gene regulation suggests that there are parallels between CdLS and Roberts syndrome (Tonkin, 2004).

Highlights

  • Normal exon-intron organization and expressed products of the gene NIPBL severed by the 5p13 breakpoint.
  • Specific embryonic expression of NIPBL transcripts.
  • Expression of NIPBL in the developing mouse.

Discussion
The involvement of Nipped-B in activating the Ubx and Cut homeobox genes may provide insights into the molecular basis of CdLS pathogenesis. Ubx suppresses limb formation in the fly abdomen by repressing Distalless (Dll), a gene required for distal limb development. The Dlx family of mammalian Dll homologs are involved in multiple developmental processes, including limb and branchial arch patterning, neurogenesis and hematopoiesis. They are expressed in the apical ectodermal ridge of the developing limb bud, which partly coordinates limb outgrowth, and also in facial primordia. Therefore, the proximodistal limb patterning defect that underlies limb reduction in CdLS and possibly the associated facial abnormalities could largely be explained by inappropriate activation of DLX genes. Mutations in fly Cut cause leg and wing abnormalities. The mouse homolog Cutl2 (Cux2) is dynamically expressed in branchial arch and limb bud progress zones, and so reduced expression of a human homolog in CdLS could also contribute to facial and limb abnormalities. The other mouse homolog, Cutl1, is widely expressed but important in lung development (Tonkin, 2004).

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Reviews

Dorsett D. (2013). What fruit flies can tell us about human birth defects. Mo Med 110: 309-313. PubMed ID: 24003648

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Date revised: 8 Jan 2016

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