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Regulatory variant in FZD6 gene contributes to nonsyndromic cleft lip and palate in an African-American family.

Cvjetkovic N, Maili L, Weymouth KS, Hashmi SS, Mulliken JB, Topczewski J, Letra A, Yuan Q, Blanton SH, Swindell EC, Hecht JT - Mol Genet Genomic Med (2015)

Bottom Line: The variant C allele segregated with NSCLP in this family, through affected and unaffected individuals, and was found in one other NSCLP African-American family.Functional assays showed that this allele creates an allele-specific protein-binding site and decreases promoter activity.We hypothesize, therefore, that alteration in FZD6 expression contributes to NSCLP in this family by perturbing the WNT signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, University of Texas Medical School at Houston Houston, Texas ; Graduate School of Biomedical Sciences, University of Texas Health Science Center Houston, Texas.

ABSTRACT
Nonsyndromic cleft lip with or without cleft palate (NSCLP) is a common birth defect affecting 135,000 newborns worldwide each year. While a multifactorial etiology has been suggested as the cause, despite decades of research, the genetic underpinnings of NSCLP remain largely unexplained. In our previous genome-wide linkage study of a large NSCLP African-American family, we identified a candidate locus at 8q21.3-24.12 (LOD = 2.98). This region contained four genes, Frizzled-6 (FZD6), Matrilin-2 (MATN2), Odd-skipped related 2 (OSR2) and Solute Carrier Family 25, Member 32 (SLC25A32). FZD6 was located under the maximum linkage peak. In this study, we sequenced the coding and noncoding regions of these genes in two affected family members, and identified a rare variant in intron 1 of FZD6 (rs138557689; c.-153 + 432A>C). The variant C allele segregated with NSCLP in this family, through affected and unaffected individuals, and was found in one other NSCLP African-American family. Functional assays showed that this allele creates an allele-specific protein-binding site and decreases promoter activity. We also observed that loss and gain of fzd6 in zebrafish contributes to craniofacial anomalies. FZD6 regulates the WNT signaling pathway, which is involved in craniofacial development, including midfacial formation and upper labial fusion. We hypothesize, therefore, that alteration in FZD6 expression contributes to NSCLP in this family by perturbing the WNT signaling pathway.

No MeSH data available.


Related in: MedlinePlus

Schematic model of WNT signaling by the FZD receptor(s). (A) In absence of WNT, β-catenin is degraded by AXIN-APC-GSK3β complex. (B) Binding of WNT to FZD receptor (plus specific other coreceptors) results in transcription of WNT target genes by the canonical pathway, mediation of tissue polarity control by noncanonical DVL-dependent pathway, and inhibition of RNA transcription of WNT target genes by noncanonical Ca2+- dependent pathway. APC, adenomatous polyposis coli; AXIN, axis inhibition protein; CAMKII, Calcium/calmodulin-dependent protein kinase II; DVL, disheveled; GSK3β, glycogen synthase kinase 3 beta; JNK, Jun N-terminal kinase; LRP5/6, low-density lipoprotein receptor-related protein 5/6; NLK, Nemo-like kinase; PIP2, Phosphatidylinositol-4,5-bisphosphate; Rho/Rac, Small GTP-binding proteins; TCF/LEF, T-cell factor/Lymphoid enhancer-binding factor; WNT, Wingless-type MMTV integration site family.
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fig04: Schematic model of WNT signaling by the FZD receptor(s). (A) In absence of WNT, β-catenin is degraded by AXIN-APC-GSK3β complex. (B) Binding of WNT to FZD receptor (plus specific other coreceptors) results in transcription of WNT target genes by the canonical pathway, mediation of tissue polarity control by noncanonical DVL-dependent pathway, and inhibition of RNA transcription of WNT target genes by noncanonical Ca2+- dependent pathway. APC, adenomatous polyposis coli; AXIN, axis inhibition protein; CAMKII, Calcium/calmodulin-dependent protein kinase II; DVL, disheveled; GSK3β, glycogen synthase kinase 3 beta; JNK, Jun N-terminal kinase; LRP5/6, low-density lipoprotein receptor-related protein 5/6; NLK, Nemo-like kinase; PIP2, Phosphatidylinositol-4,5-bisphosphate; Rho/Rac, Small GTP-binding proteins; TCF/LEF, T-cell factor/Lymphoid enhancer-binding factor; WNT, Wingless-type MMTV integration site family.

Mentions: FZD6 is part of the Frizzled family of genes that encode a group of G-coupled receptors critical for initiation of WNT signaling and is expressed in the mandible and maxilla during murine development (Tokuhara et al. 1998; Borello et al. 1999; MacDonald et al. 2009). WNT signaling is a highly controlled cellular pathway that regulates multiple functions during craniofacial development through both noncanonical and canonical/β-catenin signal transduction (Fig.4) (Logan and Nusse 2004; Moon et al. 2004; De Calisto et al. 2005; Jiang et al. 2006; Brugmann et al. 2007; Song et al. 2009; Mani et al. 2010; Mazemondet et al. 2011; Reid et al. 2011; Wang et al. 2011). It has been suggested that FZD6 represses canonical WNT signaling through the noncanonical Ca2+/CaMKII pathway by downregulating TCF/LEF-binding activity and subsequent transcription of WNT target genes (Fig.4) (Golan et al. 2004). Therefore, we hypothesize that decreased FZD6 expression could lead to dysregulation of the WNT signaling pathway, a tissue-dependent effect leading to isolated clefting. Support for this theory comes from studies showing severe craniofacial abnormalities in mice deficient for Dkk1 (Dickkopf-related protein 1), which is also a negative regulator of the WNT pathway (Mukhopadhyay et al. 2001). Additionally, a long-range murine enhancer of Myc expression causes dysregulation of Fzd6 as well as Wnt5A, Wnt9b, TCF4, Dkk1, and Lef1, all WNT pathway genes (Uslu et al. 2014). This suggests that any alteration in physiological levels of fzd6 results in abnormal craniofacial development and that fzd6 levels must be finely regulated for normal development to occur. Altogether, these findings suggest a complex role for this WNT regulator in development.


Regulatory variant in FZD6 gene contributes to nonsyndromic cleft lip and palate in an African-American family.

Cvjetkovic N, Maili L, Weymouth KS, Hashmi SS, Mulliken JB, Topczewski J, Letra A, Yuan Q, Blanton SH, Swindell EC, Hecht JT - Mol Genet Genomic Med (2015)

Schematic model of WNT signaling by the FZD receptor(s). (A) In absence of WNT, β-catenin is degraded by AXIN-APC-GSK3β complex. (B) Binding of WNT to FZD receptor (plus specific other coreceptors) results in transcription of WNT target genes by the canonical pathway, mediation of tissue polarity control by noncanonical DVL-dependent pathway, and inhibition of RNA transcription of WNT target genes by noncanonical Ca2+- dependent pathway. APC, adenomatous polyposis coli; AXIN, axis inhibition protein; CAMKII, Calcium/calmodulin-dependent protein kinase II; DVL, disheveled; GSK3β, glycogen synthase kinase 3 beta; JNK, Jun N-terminal kinase; LRP5/6, low-density lipoprotein receptor-related protein 5/6; NLK, Nemo-like kinase; PIP2, Phosphatidylinositol-4,5-bisphosphate; Rho/Rac, Small GTP-binding proteins; TCF/LEF, T-cell factor/Lymphoid enhancer-binding factor; WNT, Wingless-type MMTV integration site family.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4585452&req=5

fig04: Schematic model of WNT signaling by the FZD receptor(s). (A) In absence of WNT, β-catenin is degraded by AXIN-APC-GSK3β complex. (B) Binding of WNT to FZD receptor (plus specific other coreceptors) results in transcription of WNT target genes by the canonical pathway, mediation of tissue polarity control by noncanonical DVL-dependent pathway, and inhibition of RNA transcription of WNT target genes by noncanonical Ca2+- dependent pathway. APC, adenomatous polyposis coli; AXIN, axis inhibition protein; CAMKII, Calcium/calmodulin-dependent protein kinase II; DVL, disheveled; GSK3β, glycogen synthase kinase 3 beta; JNK, Jun N-terminal kinase; LRP5/6, low-density lipoprotein receptor-related protein 5/6; NLK, Nemo-like kinase; PIP2, Phosphatidylinositol-4,5-bisphosphate; Rho/Rac, Small GTP-binding proteins; TCF/LEF, T-cell factor/Lymphoid enhancer-binding factor; WNT, Wingless-type MMTV integration site family.
Mentions: FZD6 is part of the Frizzled family of genes that encode a group of G-coupled receptors critical for initiation of WNT signaling and is expressed in the mandible and maxilla during murine development (Tokuhara et al. 1998; Borello et al. 1999; MacDonald et al. 2009). WNT signaling is a highly controlled cellular pathway that regulates multiple functions during craniofacial development through both noncanonical and canonical/β-catenin signal transduction (Fig.4) (Logan and Nusse 2004; Moon et al. 2004; De Calisto et al. 2005; Jiang et al. 2006; Brugmann et al. 2007; Song et al. 2009; Mani et al. 2010; Mazemondet et al. 2011; Reid et al. 2011; Wang et al. 2011). It has been suggested that FZD6 represses canonical WNT signaling through the noncanonical Ca2+/CaMKII pathway by downregulating TCF/LEF-binding activity and subsequent transcription of WNT target genes (Fig.4) (Golan et al. 2004). Therefore, we hypothesize that decreased FZD6 expression could lead to dysregulation of the WNT signaling pathway, a tissue-dependent effect leading to isolated clefting. Support for this theory comes from studies showing severe craniofacial abnormalities in mice deficient for Dkk1 (Dickkopf-related protein 1), which is also a negative regulator of the WNT pathway (Mukhopadhyay et al. 2001). Additionally, a long-range murine enhancer of Myc expression causes dysregulation of Fzd6 as well as Wnt5A, Wnt9b, TCF4, Dkk1, and Lef1, all WNT pathway genes (Uslu et al. 2014). This suggests that any alteration in physiological levels of fzd6 results in abnormal craniofacial development and that fzd6 levels must be finely regulated for normal development to occur. Altogether, these findings suggest a complex role for this WNT regulator in development.

Bottom Line: The variant C allele segregated with NSCLP in this family, through affected and unaffected individuals, and was found in one other NSCLP African-American family.Functional assays showed that this allele creates an allele-specific protein-binding site and decreases promoter activity.We hypothesize, therefore, that alteration in FZD6 expression contributes to NSCLP in this family by perturbing the WNT signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, University of Texas Medical School at Houston Houston, Texas ; Graduate School of Biomedical Sciences, University of Texas Health Science Center Houston, Texas.

ABSTRACT
Nonsyndromic cleft lip with or without cleft palate (NSCLP) is a common birth defect affecting 135,000 newborns worldwide each year. While a multifactorial etiology has been suggested as the cause, despite decades of research, the genetic underpinnings of NSCLP remain largely unexplained. In our previous genome-wide linkage study of a large NSCLP African-American family, we identified a candidate locus at 8q21.3-24.12 (LOD = 2.98). This region contained four genes, Frizzled-6 (FZD6), Matrilin-2 (MATN2), Odd-skipped related 2 (OSR2) and Solute Carrier Family 25, Member 32 (SLC25A32). FZD6 was located under the maximum linkage peak. In this study, we sequenced the coding and noncoding regions of these genes in two affected family members, and identified a rare variant in intron 1 of FZD6 (rs138557689; c.-153 + 432A>C). The variant C allele segregated with NSCLP in this family, through affected and unaffected individuals, and was found in one other NSCLP African-American family. Functional assays showed that this allele creates an allele-specific protein-binding site and decreases promoter activity. We also observed that loss and gain of fzd6 in zebrafish contributes to craniofacial anomalies. FZD6 regulates the WNT signaling pathway, which is involved in craniofacial development, including midfacial formation and upper labial fusion. We hypothesize, therefore, that alteration in FZD6 expression contributes to NSCLP in this family by perturbing the WNT signaling pathway.

No MeSH data available.


Related in: MedlinePlus