scholarly journals Functional SNPs within the Intron 1 of the PROP1 Gene Contribute to Combined Growth Hormone Deficiency (CPHD)

2012 ◽  
Vol 97 (9) ◽  
pp. E1791-E1797 ◽  
Author(s):  
Michela Godi ◽  
Simona Mellone ◽  
Luigi Tiradani ◽  
Rita Marabese ◽  
Claudio Bardelli ◽  
...  
Keyword(s):  
Regulatory Element ◽  
Hormone Deficiency ◽  
Luciferase Reporter ◽  
Pituitary Hormone ◽  
Mcf7 Cells ◽  
Luciferase Reporter Gene ◽  
Risk Alleles ◽  
Intron 1 ◽  
Sporadic Cases ◽  
Prop1 Gene

Context: Mutations within the PROP1 gene represent one of the main causes of familial combined pituitary hormone deficiency (CPHD). However, most of the cases are sporadic with an unknown genetic cause. Objective: The aim of this study was the search for low penetrance variations within and around a conserved regulatory element in the intron 1 of PROP1, contributing to a multifactorial form of the disease in sporadic patients. Methods and Patients: A fragment of 570 bp encompassing the conserved region was sequenced in 107 CPHD patients and 294 controls, and an association study was performed with the four identified variants, namely c.109+435G>A (rs73346254), c.109+463C>T (rs4498267), c.109+768C>G (rs4431364), and c.109+915_917ins/delTAG (rs148607624). The functional role of the associated polymorphisms was evaluated by luciferase reporter gene expression analyses and EMSA. Results: A statistically significant increased frequency was observed in the patients for rs73346254A (P = 5 × 10−4) and rs148607624delTAG (P = 0.01) alleles. Among all the possible allele combinations, only the haplotype bearing both risk alleles showed a significantly higher frequency in the patients vs. controls (P = 4.7 × 10−4) and conferred a carrier risk of 4.19 (P = 1.2 × 10−4). This haplotype determined a significant decrease of the luciferase activity in comparison with a basal promoter and the other allelic combinations in GH4C and MCF7 cells (P = 4.6 × 10−6; P = 5.5 × 10−4, respectively). The EMSA showed a differential affinity for nuclear proteins for the alternative alleles of the two associated variations. Conclusions: Variations with a functional significance conferring susceptibility to CPHD have been identified in the PROP1 gene, indicating a multifactorial origin of this disorder in sporadic cases.

scholarly journals The PROP1 2-Base Pair Deletion Is a Common Cause of Combined Pituitary Hormone Deficiency1

1998 ◽  
Vol 83 (9) ◽  
pp. 3346-3349 ◽  
Author(s):  
Joy D. Cogan ◽  
Wei Wu ◽  
John A. Phillips ◽  
Ivo J. P. Arnhold ◽  
Ana Agapito ◽  
...  
Keyword(s):  
Human Subjects ◽  
Polymorphic Marker ◽  
Hormone Deficiency ◽  
Pituitary Hormone ◽  
Degree Relative ◽  
Exon 2 ◽  
Base Pair Deletion ◽  
Stimulation Tests ◽  
Sporadic Cases ◽  
Prop1 Gene

Combined pituitary hormone deficiency (CPHD) has an incidence of approximately 1 in 8000 births. Although the proportion of familial CPHD cases is unknown, about 10% have an affected first degree relative. We have recently reported three mutations in the PROP1 gene that cause CPHD in human subjects. We report here the frequency of one of these mutations, a 301–302delAG deletion in exon 2 of PROP1, in 10 independently ascertained CPHD kindreds and 21 sporadic cases of CPHD from 8 different countries. Our results show that 55% (11 of 20) of PROP1 alleles have the 301–302delAG deletion in familial CPHD cases. Interestingly, although only 12% (5 of 42) of the PROP1 alleles of our 21 sporadic cases were 301–302delAG, the frequency of this allele (in 20 of 21 of the sporadic subjects given TRH stimulation tests) was 50% (3 of 6) and 0% (0 of 34) in the CPHD cases with pituitary and hypothalamic defects, respectively. Using whole genome radiation hybrid analysis, we localized the PROP1 gene to the distal end of chromosome 5q and identified a tightly linked polymorphic marker, D5S408, which can be used in segregation studies. Analysis of this marker in affected subjects with the 301–302delAG deletion suggests that rather than being inherited from a common founder, the 301–302delAG may be a recurring mutation.

A mutation of the β-domain in POU1F1 causes pituitary deficiency due to dominant PIT-1β expression

2021 ◽  
Author(s):  
Shigeru Suzuki ◽  
Kumihiro Matsuo ◽  
Yoshiya Ito ◽  
Atsushi Kobayashi ◽  
Takahide Kokumai ◽  
...  
Keyword(s):  
Hormone Deficiency ◽  
Luciferase Reporter ◽  
Heterozygous Mutation ◽  
Pituitary Hormone ◽  
Pituitary Cells ◽  
Transactivation Domain ◽  
Exon 2 ◽  
Pituitary Development ◽  
Long Term Follow Up ◽  
Alternatively Spliced

Background: POU1F1 encodes both PIT-1α, which plays pivotal roles in pituitary development and GH, PRL and TSHB expression, and the alternatively spliced isoform PIT-1β, which contains an insertion of 26-amino acids (β-domain) in the transactivation domain of PIT-1α due to the use of an alternative splice acceptor at the end of the first intron. PIT-1β is expressed at much lower levels than PIT-1α and represses endogenous PIT-1α transcriptional activity. Although POU1F1 mutations lead to combined pituitary hormone deficiency (CPHD), no patients with β-domain mutations have been reported. Results: Here, we report that a three-generation family exhibited different degrees of CPHD, including growth hormone deficiency with intrafamilial variability of prolactin/TSH insufficiency and unexpected prolactinoma occurrence. The CPHD was due to a novel POU1F1 heterozygous variant (c.143-69T>G) in intron 1 of PIT-1α (RefSeq number NM_000306) or as c.152T>G (p.Ile51Ser) in exon 2 of PIT-1β (NM_001122757). Gene splicing experiments showed that this mutation yielded the PIT-1β transcript without other transcripts. Lymphocyte PIT-1β mRNA expression was significantly higher in the patients with the heterozygous mutation than a control. A luciferase reporter assay revealed that the PIT-1β-Ile51Ser mutant repressed PIT-1α and abolished transactivation capacity for the rat prolactin promoter in GH3 pituitary cells. Conclusions: We describe, for the first time, that PIT-1β mutation can cause CPHD through a novel genetic mechanism, such as PIT-1β overexpression, and that POU1F1 mutation might be associated with a prolactinoma. Analysis of new patients and long-term follow-up are needed to clarify the characteristics of PIT-1β mutations.

Chromatin Conformation and a Distal Regulatory Element Activate the Human Erythroid Ankyrin-1 Promoter.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 803-803
Author(s):  
Ashley N. Owen ◽  
Robert I. Liem ◽  
Andre M. Pilon ◽  
Patrick G. Gallagher ◽  
David M. Bodine
Keyword(s):  
Reporter Gene ◽  
Regulatory Element ◽  
K562 Cells ◽  
Regulatory Elements ◽  
Dnase I ◽  
Luciferase Reporter ◽  
Erythroid Cells ◽  
Luciferase Reporter Gene ◽  
Human And Mouse ◽  
In Erythroid Cells

Abstract Ankyrin forms the bridge between the spectrin/actin network of the erythrocyte membrane skeleton and the red cell membrane by binding to both β-spectrin and band 3. The erythrocyte ankyrin promoter (Ank-1E) is active only in erythroid cells, while two other Ank-1 promoters located 20 kb downstream and 40 kb upstream of Ank-1E are active in the cerebellum and muscle cells respectively. We have been studying the mechanism by which the Ank-1E promoter becomes active in erythroid cells by studying the cis acting regulatory elements and the chromatin structure of the Ank-1 promoter region. We have previously shown that the sequences between −296 and −15 of the Ank-1E promoter are fully sufficient for erythroid specific, copy number dependent uniform expression of reporter genes in transgenic mice. We have also mapped a DNase I Hypersensitive site (5′HS) between −300 and −100 of the human and mouse Ank-1E promoters in human K562 and mouse fetal liver cells. Both the mouse and human 5′HS are capable of preventing the silencing of a β-globin/GFP reporter gene in K562 cells, establishing that they function as barrier elements. Consistent with this observation, the human and mouse 5′HS are hyperacetylated in erythroid cells. The chromatin 10 kb 5′ to the 5′HS is DNase I resistant (associated with inactive chromatin) in human and mouse erythroid and non-erythroid cells. Approximately 6 kb 3′ to 5′HS are two adjacent HS (3′HS1, 3′HS2). Beyond 3′HS2 the chromatin is also DNase I resistant in both human and mouse erythroid and non-erythroid cells. Between 5′HS and 3′HS1 the 6kb region is DNase I sensitive (active) in erythroid cells but not in other cell types. We hypothesized that this 6 kb region contains regulatory elements that activate the Ank-1E promoter. To screen for regulatory elements we isolated overlapping segments of a 10 kb region extending from 2 kb upstream of 5′HS to 2 kb downstream of 3′HS2. We inserted these fragments into a plasmid vector containing the Ank-1E promoter linked to a luciferase reporter gene and transfected these constructs into K562 cells. A single region up regulated Ank-1E/luciferase expression. This region mapped to a 211bp segment that included 3′HS1, but did not include 3′HS2. A fragment containing only 3′HS2 did not up regulate an Ank-1E/luciferase reporter gene, but 3′HS2 was capable of preventing the silencing of a β-globin/Green Fluorescent Protein reporter gene in K562 cells, demonstrating barrier activity. The region around 3′HS1 and 2 was also a site of histone hyperacetylation. The sequence of the 211 bp fragment containing 3′HS1 does not contain consensus sequences for any known erythroid-specific transcription factors, but does contain potential binding sites fro Sp1, AP-1 and E-box binding proteins. Using the Chromatin Conformation Capture assay we demonstrated that 5′HS and 3′HS1 and 2 are in close proximity in K562 chromatin, but are not closely associated in chromatin from other cell types. We propose that an erythroid-specific chromatin loop brings 3′HS1 and 2 into proximity with 5′HS, adjacent to the Ank-1E promoter. This interaction translocates the positive regulatory element in 3′HS1 to the Ank-1E promoter allowing the Ank-1E promoter to become active in erythroid cells.

The c.301_302delAG PROP1 gene mutation in Romanian patients with multiple pituitary hormone deficiency

2015 ◽  
Vol 28 (9-10) ◽  
Author(s):  
Cecilia Lazea ◽  
Paula Grigorescu-Sido ◽  
Radu Popp ◽  
Marie Legendre ◽  
Serge Amselem ◽  
...  
Keyword(s):  
High Frequency ◽  
Bone Age ◽  
Hormone Deficiency ◽  
Pituitary Hormone ◽  
Homozygous State ◽  
Pituitary Hormone Deficiency ◽  
Homozygous Genotype ◽  
Familial Form ◽  
History Of ◽  
Prop1 Gene

AbstractTo establish the frequency of the c.301_302 delAG mutation of theSomatic assessment, hormonal test, bone age, magnetic resonance imaging of the pituitary gland, and molecular diagnosis were performed in 26 patients with MPHD (7 patients with familial form of MPHD and 19 patients with sporadic form of MPHD).The c.301_302delAG mutation was detected in the homozygous state in 10 patients belonging to 5 unrelated families (7 patients with familial history of MPHD and 3 patients with sporadic form of MPHD). Those 10 patients presented variable pituitary hormone deficiency and pituitary morphology.The c.301_302delAG homozygous genotype had a high frequency of 38% (10/26), reaching 100% (7/7) in group with familial cases of MPHD and 16% (3/19) in group with sporadic forms of MPHD.

scholarly journals Molecular analysis of the PROP1 and HESX1 genes in patients with septo-optic dysplasia and/or pituitary hormone deficiency

2010 ◽  
Vol 54 (5) ◽  
pp. 482-487 ◽  
Author(s):  
Juliana B. Cruz ◽  
Vania S. Nunes ◽  
Sueli A. Clara ◽  
Denise Perone ◽  
Peter Kopp ◽  
...  
Keyword(s):  
Gene Mutations ◽  
Genetic Alterations ◽  
Hormone Deficiency ◽  
Pituitary Hormone ◽  
Pituitary Hormone Deficiency ◽  
Exon 1 ◽  
Genetic Mechanisms ◽  
Direct Sequence Analysis ◽  
Prop1 Gene ◽  
Pathogenic Process

OBJECTIVE: The present study aimed at evaluating the PROP1 and HESX1 genes in a group of patients with septo-optic dysplasia (SOD) and pituitary hormone deficiency (combined - CPHD; isolated GH deficiency - GHD). Eleven patients with a clinical and biochemical presentation consistent with CPHD, GHD or SOD were evaluated. SUBJECTS AND METHODS: In all patients, the HESX1 gene was analyzed by direct sequence analysis and in cases of CPHD the PROP1 gene was also sequenced. RESULTS: A polymorphism (1772 A > G; N125S) was identified in a patient with SOD. We found three patients carrying the allelic variants 27 T > C; A9A and 59 A > G; N20S in exon 1 of the PROP1 gene. Mutations in the PROP1 and HESX1 genes were not identified in these patients with sporadic GHD, CPHD and SOD. CONCLUSION: Genetic alterations in one or several other genes, or non-genetic mechanisms, must be implicated in the pathogenic process.

scholarly journals PROP1 gene analysis in Portuguese patients with combined pituitary hormone deficiency

2006 ◽  
Vol 65 (4) ◽  
pp. 479-485 ◽  
Author(s):  
Manuel C. Lemos ◽  
Leonor Gomes ◽  
Margarida Bastos ◽  
Valeriano Leite ◽  
Edward Limbert ◽  
...  
Keyword(s):  
Gene Analysis ◽  
Hormone Deficiency ◽  
Pituitary Hormone ◽  
Pituitary Hormone Deficiency ◽  
Combined Pituitary Hormone Deficiency ◽  
Prop1 Gene

scholarly journals Insulin activates the rat sterol-regulatory-element-binding protein 1c (SREBP-1c) promoter through the combinatorial actions of SREBP, LXR, Sp-1 and NF-Y cis-acting elements

2004 ◽  
Vol 385 (1) ◽  
pp. 207-216 ◽  
Author(s):  
Lauren M. CAGEN ◽  
Xiong DENG ◽  
Henry G. WILCOX ◽  
Edwards A. PARK ◽  
Rajendra RAGHOW ◽  
...  
Keyword(s):  
Binding Protein ◽  
Regulatory Element ◽  
Ectopic Expression ◽  
Rat Hepatocytes ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Cis Acting ◽  
Element Binding Protein ◽  
Sterol Regulatory Element

The enhanced synthesis of fatty acids in the liver and adipose tissue in response to insulin is critically dependent on the transcription factor SREBP-1c (sterol-regulatory-element-binding protein 1c). Insulin increases the expression of the SREBP-1c gene in intact liver and in hepatocytes cultured in vitro. To learn the mechanism of this stimulation, we analysed the activation of the rat SREBP-1c promoter and its truncated or mutated congeners driving a luciferase reporter gene in transiently transfected rat hepatocytes. The rat SREBP-1c promoter contains binding sites for LXR (liver X receptor), Sp1, NF-Y (nuclear factor-Y) and SREBP itself. We have found that each of these sites is required for the full stimulatory response of the SREBP-1c promoter to insulin. Mutation of either the putative LXREs (LXR response elements) or the SRE (sterol response element) in the proximal SREBP-1c promoter reduced the stimulatory effect of insulin by about 50%. Insulin and the LXR agonist TO901317 increased the association of SREBP-1 with the SREBP-1c promoter. Ectopic expression of LXRα or SREBP-1c increased activity of the SREBP-1c promoter, and this effect is further enhanced by insulin. The Sp1 and NF-Y sites adjacent to the SRE are also required for full activation of the SREBP-1c promoter by insulin. We propose that the combined actions of the SRE, LXREs, Sp1 and NF-Y elements constitute an insulin-responsive cis-acting unit of the SREBP-1c gene in the liver.

scholarly journals The R271W mutant form of Pit-1 does not act as a dominant inhibitor of Pit-1 action to activate the promoters of GH and prolactin genes

2003 ◽  
pp. 619-625 ◽  
Author(s):  
M Kishimoto ◽  
Y Okimura ◽  
M Fumoto ◽  
G Iguchi ◽  
K Iida ◽  
...  
Keyword(s):  
Reporter Gene ◽  
Dominant Negative ◽  
Hormone Deficiency ◽  
Luciferase Reporter ◽  
Dominant Negative Effect ◽  
Pituitary Hormone ◽  
Mutant Form ◽  
Wild Type ◽  
Expression Study ◽  
Negative Effect

OBJECTIVE: Genetic abnormalities of the pituitary specific transcription factor, Pit-1, have been reported in several patients with GH, prolactin (PRL) and TSH deficiencies. The most common is a mutation altering an arginine to a tryptophan in codon 271 (R271W) in one allele of the Pit-1 gene. According to the previous in vitro expression study, R271W acted as a dominant negative inhibitor of the wild type to activate the GH promoter. However, healthy carriers with this mutation, who should be affected by the dominant negative effect of R271W, have also been reported. The aim of this study was to clarify in more detail the function of this mutant form of Pit-1. METHODS: Transcriptional activity of R271W for the expression of Pit-1-associated genes was investigated in COS7 cells with the aid of transient transfection assays. The 1.8 kb rat GH, 0.6 kb rat PRL or 1.9 kb rat PRL 5'-flanking regions were inserted upstream of the luciferase reporter gene and were used for functional analysis of R271W. Another reporter gene containing seven Pit-1 responsive elements was also used. The same experiments were also performed using JEG3 and CHO cells. RESULTS: We could not confirm the dominant negative effect of R271W on wild type Pit-1. Furthermore, our expression study revealed that R271W could activate the promoters of GH and PRL genes to levels similar to the wild type. CONCLUSION: Taken together with the evidence that phenotypically normal cases have been reported with this mutation, our results deny the relationship between R271W and combined pituitary hormone deficiency.

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