On the molecular etiology of Cornelia de Lange syndrome

Abstract

Cornelia de Lange syndrome (CdLS) is genetically heterogeneous and is usually sporadic, occurring approximately once per 10,000 births. CdLS individuals display diverse and variable deficits in growth, mental development, limbs, and organs. In the past few years it has been shown that CdLS is caused by gene mutations affecting proteins involved in sister chromatid cohesion. Studies in model organisms, and more recently in human cells, have revealed, somewhat unexpectedly, that the developmental deficits in CdLS likely arise from changes in gene expression. The mechanisms by which cohesion factors regulate gene expression remain to be elucidated, but current data suggest that they likely regulate transcription in multiple ways.

Fig. 1. Phenotypic spectrum in CdLS

Characteristic facial features in two individuals with CdLS and NIPBL mutations (a and b) and in two children with SMC1A mutations (c and d). Note the arched eyebrows with synophrys, long eyelashes, ptosis (more noticeable in `a' and `b'), the short upturned nose and long philtrum with thin upper lip. These features are still present in the SMC1A mutated individuals, but to a more subtle degree. The variability of the phenotype in CdLS is highlighted by the range of involvement of the upper limbs as demonstrated in e-j. In `e' the more severe end of the spectrum of upper limb involvement is depicted with severe ulnar hypoplasia of the forearm with only a single digit and underdevelopment of almost all boney structures. f-h depicts variable forms of oligodactyly and i-j demonstrates the milder end of the spectrum with small hands, 5^th finger clinodactyly and proximally placed thumbs.

Fig. 2. The cohesin complex and regulatory factors

(Upper diagram) Cohesin consists of two SMC (structural maintenance of chromosome) proteins, Smc1 and Smc3, the kleisin Rad21 (Mcd1, Scc1) and Stromalin (Stag2, SA, Scc3).- Each SMC protein folds back on themselves at the hinge region to form long antiparallel coiled-coil arms. The N and C termini of each SMC protein come together to form an ABC-type ATPase head domain. The N and C termini of Rad21 interact with the head domains of Smc3 and Smc1, respectively. A kink in the Smc3 arm creates an open ring-like structure with an internal diameter of ~35 nm. (Lower diagram) A leading hypothesis is that cohesin mediates sister chromatid cohesion by encircling the two sister chromatids. NIPBL (Nipped-B, Scc2, Mis4) interacts with the Mau-2 (Scc4) protein and is required for binding of cohesin to chromosomes. The interactions of the NIPBL/Mau-2 complex with cohesin shown in the diagram are speculative. The Pds5 protein interacts with the hinge domain of Smc1 and Rad21 (not shown), and is required for sister chromatid cohesion, but not binding of cohesin to chromosomes.

Fig. 3. Nipped-B and cohesin associate with the transcribed Abdominal-B (Abd-B) gene in the Drosophila bithorax complex (BX-C)

The BX-C contains the Ultrabithorax (Ubx), abdominal-A (abd-A) and Abd-B homeobox genes (transcribed from right to left) that control segmental identity and limb development. Abd-B is highly transcribed in Sg4 cells of embryonic origin, but not in MLDmBG3 (BG3) cells derived larval central nervous system., The upper three tracks show the histone H3 lysine 27 trimethylation (H3K27Me), a mark of Polycomb group (PcG) silencing, across the BX-C in Sg4 cells, and the binding of RNA polymerase II (PolII) and Nipped-B, the Drosophila NIPBL ortholog. The pattern of cohesin binding (not shown) is identical to that of Nipped-B. The middle tracks show the locations of know insulator/boundary elements in the BX-C, and binding sites for the CCTC-binding factor (CTCF). The binding of PolII and Nipped-B is restricted to the region between a known insulator (Fab-7, not labeled), and a CTCF insulator protein binding site near the Abd-B distal promoter, which also demarcate the boundaries between the active Abd-B region and the flanking silenced domains. The two bottom tracks show the lack of significant PolII or cohesin binding to the inactive BX-C in BG3 cells.

Fig. 4. Cohesin contributes to insulation by CTCF at the imprinted insulator in the human H19-Igf2 locus.

The H19-Igf2 locus contains a CTCF-dependent insulator that is regulated by imprinting. In the maternally-inherited chromosome (Maternal), CTCF binds the insulator, preventing an enhancer located downstream of H19 from activating Igf2, and forcing it to active H19 (transcribed from right to left). In the paternally-inherited chromosome (Paternal), the insulator DNA is methylated (Me), which prevents binding of CTCF. The insulator no longer functions, allowing the enhancer to activate Igf2 (transcribed from right to left) instead of H19. Cohesin co-localizes with CTCF at many sites in mammalian cells, including the H19-Igf2 insulator.,, The two tracks above the gene map show the binding of the Rad21 cohesin subunit and CTCF to the H19-Igf2 locus in HeLa cells. Knockdown of either CTCF or Rad21 in HeLa cells simultaneously increases Igf2 transcripts and decreases H19 transcripts, consistent with a reduction in insulator activity.