A humanized IKBKAP transgenic mouse models a tissue-specific human splicing defect

Hims, Matthew M.; Shetty, Ranjit S.; Pickel, James; Mull, James; Leyne, Maire; Liu, Lijuan; Gusella, James F.; Slaugenhaupt, Susan A.

doi:10.1016/j.ygeno.2007.05.012

Cited by 55 publications

(82 citation statements)

References 25 publications

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“…The day of vaginal-plug discovery was designated 0.5 dpc. The transgenic mouse lines that carry human IKBKAP transgenes were generated as previously reported (23). The mice used for this study were housed in the animal facility of Massachusetts General Hospital (Boston, MA), provided with constant access to a standard diet of food and water, and maintained on a 12-hour light/dark cycle, and all experimental protocols were approved by the Subcommittee on Research Animal Care at the Massachusetts General Hospital.…”

Section: Animalsmentioning

confidence: 99%

“…Recently, we described the creation of transgenic mouse lines that carry the normal human IKBKAP gene and the FD IVS20ϩ6TϾC mutation. All of these lines are phenotypically normal, and the IVS20ϩ6TϾC transgenics missplice IKBKAP in a tissue-specific manner that mimics what is seen in FD patients (23). To test whether embryonic lethality caused by ablation of mouse Ikbkap could be rescued by human IKAP, we crossed two transgenic lines, one carrying the normal IKBKAP gene and one with the IVS20ϩ6TϾC mutation, with Ikbkap ϩ/Ϫ mice in order to introduce the human transgenes into the Ikbkap knockout mouse line.…”

Section: Fig 4 Appearance Of Ikbkapmentioning

confidence: 99%

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Loss of Mouse Ikbkap, a Subunit of Elongator, Leads to Transcriptional Deficits and Embryonic Lethality That Can Be Rescued by Human IKBKAP

Chen

Hims

Shetty

et al. 2009

Molecular and Cellular Biology

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107

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Familial dysautonomia (FD), a devastating hereditary sensory and autonomic neuropathy, results from an intronic mutation in the IKBKAP gene that disrupts normal mRNA splicing and leads to tissue-specific reduction of IKBKAP protein (IKAP) in the nervous system. To better understand the roles of IKAP in vivo, an Ikbkap knockout mouse model was created. Results from our study show that ablating Ikbkap leads to embryonic lethality, with no homozygous Ikbkap knockout (Ikbkap ؊/؊ ) embryos surviving beyond 12.5 days postcoitum. Morphological analyses of the Ikbkap ؊/؊ conceptus at different stages revealed abnormalities in both the visceral yolk sac and the embryo, including stunted extraembryonic blood vessel formation, delayed entry into midgastrulation, disoriented dorsal primitive neural alignment, and failure to establish the embryonic vascular system. Further, we demonstrate downregulation of several genes that are important for neurulation and vascular development in the Ikbkap ؊/؊ embryos and show that this correlates with a defect in transcriptional elongation-coupled histone acetylation. Finally, we show that the embryonic lethality resulting from Ikbkap ablation can be rescued by a human IKBKAP transgene. For the first time, we demonstrate that IKAP is crucial for both vascular and neural development during embryogenesis and that protein function is conserved between mouse and human.IKBKAP (encoding IB kinase-associated protein, also called Elongator protein 1) is the gene mutated in hereditary sensory and autonomic neuropathy type III, or familial dysautonomia (FD). All FD patients carry at least one copy of a splicing mutation in IKBKAP, which causes aberrant exon skipping and subsequent tissue-specific reduction of protein expression in FD patients (1,41,42). The IKBKAP gene is highly conserved across species, with the human and mouse proteins (IKAP and Ikap, respectively) sharing more than 80% amino acid homology (12). The IKBKAP protein, IKAP, was first reported to act as a scaffolding protein for the IB kinase complex (11). Recent studies, however, have shown that IKAP does not play a role in NF-B (nuclear factor B) signaling, but rather, it is a subunit of the human Elongator complex, which is important for efficient transcriptional elongation (19,28,36).FD (or Riley-Day syndrome) is one of the best known recessive hereditary neuropathies, with an extremely high carrier frequency in the Ashkenazi Jewish population, which ranges from 1 in 17 to 1 in 28 depending on the country of origin (29,33,42). Clinical characteristics of FD include diminished tear secretion, dysphagia, esophageal and gastric dysmotility, gastroesophageal reflux, spinal curvature, postural hypotension, blotching, excessive sweating, and decreased deep-tendon reflexes (2). Fatality in FD patients is high, and only half survive to 40 years of age. Clinical reports have shown that the failure of autonomic function is one of the major causes of death (21). To date, three FD-causing mutations have been identified in the IKBKAP gene: a...

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Section: Animalsmentioning

confidence: 99%

Section: Fig 4 Appearance Of Ikbkapmentioning

confidence: 99%

Loss of Mouse Ikbkap, a Subunit of Elongator, Leads to Transcriptional Deficits and Embryonic Lethality That Can Be Rescued by Human IKBKAP

Chen

Hims

Shetty

et al. 2009

Molecular and Cellular Biology

Self Cite

107

119

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“…There is no evidence that the mutant transcript yields IKAP protein, and therefore, FD can be considered as a partial loss of IKBKAP function disease. Interestingly, the extent of IKBKAP missplicing in FD patients shows considerable tissue-specific differences [32,40] that may underlie the tissue-specific clinical symptoms of the disease.…”

Section: Introductionmentioning

confidence: 99%

“…It has been shown that mice null for IKBKAP are developmentally arrested at midgastrulation resulting in early embryonic lethality [47]. The generation of bacterial artificial chromosome (BAC) transgenic mice carrying the human BAC with the FD disease-causing mutation has been used to model tissue-specific splicing of the IKBKAP gene [40]. However, introducing the human disease-specific BAC onto an IKBKAP-null mouse background leads to a complete rescue of embryonic lethality without any signs of disease in the resulting mouse [47].…”

Section: Introductionmentioning

confidence: 99%

Modelling familial dysautonomia in human induced pluripotent stem cells

Lee

Studer

2011

Phil. Trans. R. Soc. B

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Induced pluripotent stem (iPS) cells have considerable promise as a novel tool for modelling human disease and for drug discovery. While the generation of disease-specific iPS cells has become routine, realizing the potential of iPS cells in disease modelling poses challenges at multiple fronts. Such challenges include selecting a suitable disease target, directing the fate of iPS cells into symptom-relevant cell populations, identifying disease-related phenotypes and showing reversibility of such phenotypes using genetic or pharmacological approaches. Finally, the system needs to be scalable for use in modern drug discovery. Here, we will discuss these points in the context of modelling familial dysautonomia (FD, Riley–Day syndrome, hereditary sensory and autonomic neuropathy III (HSAN-III)), a rare genetic disorder in the peripheral nervous system. We have demonstrated three disease-specific phenotypes in FD-iPS-derived cells that can be partially rescued by treating cells with the plant hormone kinetin. Here, we will discuss how to use FD-iPS cells further in high throughput drug discovery assays, in modelling disease severity and in performing mechanistic studies aimed at understanding disease pathogenesis. FD is a rare disease but represents an important testing ground for exploring the potential of iPS cell technology in modelling and treating human disease.

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“…As a result, a 3Ј-end-truncated mRNA transcript of IKBKAP is expressed at variable levels in different tissues together with full-length IKBKAP (Slaugenhaupt et al, 2001;Anderson et al, 2001). The pathogenic result of the IKBKAP splice mutation is a tissue-specific reduction of wild-type IKAP protein below a tolerable threshold in the sensory and autonomous nervous systems (Slaugenhaupt et al, 2001;Cuajungco et al, 2003;Hims et al, 2007). It is not known whether the truncated IKAP transcript resulting from the exon skipping is degraded or whether it is translated into a C-terminally truncated IKAP protein termed here as FD-IKAP.…”

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confidence: 99%

IKAP localizes to membrane ruffles with filamin A and regulates actin cytoskeleton organization and cell migration

Johansen

Naumanen

Knudsen

et al. 2008

Journal of Cell Science

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Loss-of-function mutations in the IKBKAP gene, which encodes IKAP (ELP1), cause familial dysautonomia (FD), with defective neuronal development and maintenance. Molecular mechanisms leading to FD are poorly understood. We demonstrate that various RNA-interference-based depletions of IKAP lead to defective adhesion and migration in several cell types, including rat primary neurons. The defects could be rescued by reintroduction of wild-type IKAP but not by FD-IKAP, a truncated form of IKAP constructed according to the mutation found in the majority of FD patients. Cytosolic IKAP co-purified with proteins involved in cell migration, including filamin A, which is also involved in neuronal migration. Immunostaining of IKAP and filamin A revealed a distinct co-localization of these two proteins in membrane ruffles. Depletion of IKAP resulted in a significant decrease in filamin A localization in membrane ruffles and defective actin cytoskeleton organization, which both could be rescued by the expression of wild-type IKAP but not by FD-IKAP. No downregulation in the protein levels of paxillin or beclin 1, which were recently described as specific transcriptional targets of IKAP, was detected. These results provide evidence for the role of the cytosolic interactions of IKAP in cell adhesion and migration, and support the notion that cell-motility deficiencies could contribute to FD.

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A humanized IKBKAP transgenic mouse models a tissue-specific human splicing defect

Cited by 55 publications

References 25 publications

Loss of Mouse Ikbkap, a Subunit of Elongator, Leads to Transcriptional Deficits and Embryonic Lethality That Can Be Rescued by Human IKBKAP

Loss of Mouse Ikbkap, a Subunit of Elongator, Leads to Transcriptional Deficits and Embryonic Lethality That Can Be Rescued by Human IKBKAP

Modelling familial dysautonomia in human induced pluripotent stem cells

IKAP localizes to membrane ruffles with filamin A and regulates actin cytoskeleton organization and cell migration

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