Limits...
ZBTB20 is required for anterior pituitary development and lactotrope specification.

Cao D, Ma X, Cai J, Luan J, Liu AJ, Yang R, Cao Y, Zhu X, Zhang H, Chen YX, Shi Y, Shi GX, Zou D, Cao X, Grusby MJ, Xie Z, Zhang WJ - Nat Commun (2016)

Bottom Line: Disruption of Zbtb20 leads to anterior pituitary hypoplasia, hypopituitary dwarfism and a complete loss of mature lactotropes.Furthermore, endogenous ZBTB20 protein binds to Prl promoter, and its knockdown decreases PRL expression and secretion in a lactotrope cell line MMQ.In addition, ZBTB20 overexpression enhances the transcriptional activity of Prl promoter in vitro.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathophysiology, Second Military Medical University, 800 Xiangyin Road, Shanghai 200433, China.

ABSTRACT
The anterior pituitary harbours five distinct hormone-producing cell types, and their cellular differentiation is a highly regulated and coordinated process. Here we show that ZBTB20 is essential for anterior pituitary development and lactotrope specification in mice. In anterior pituitary, ZBTB20 is highly expressed by all the mature endocrine cell types, and to some less extent by somatolactotropes, the precursors of prolactin (PRL)-producing lactotropes. Disruption of Zbtb20 leads to anterior pituitary hypoplasia, hypopituitary dwarfism and a complete loss of mature lactotropes. In ZBTB20- mice, although lactotrope lineage commitment is normally initiated, somatolactotropes exhibit profound defects in lineage specification and expansion. Furthermore, endogenous ZBTB20 protein binds to Prl promoter, and its knockdown decreases PRL expression and secretion in a lactotrope cell line MMQ. In addition, ZBTB20 overexpression enhances the transcriptional activity of Prl promoter in vitro. In conclusion, our findings point to ZBTB20 as a critical regulator of anterior pituitary development and lactotrope specification.

No MeSH data available.


Related in: MedlinePlus

Impaired development of endocrine cells in the anterior pituitary of Zbtb20- mice.(a) All five types of cell lineages in anterior pituitary are detected by immunohistochemical staining using GH, PRL, TSHβ, ACTH and LHβ antibodies in 21-day-old Zbtb20+/+ and Zbtb20−/− littermate. PRL-positive cells are absent in  mice, but other cell lineages have no apparent changes. (b) Immunohistochemical staining of Drd2 in 21-day-old pituitary sections also shows a loss of positive cells in the  mice. Insets show the whole coronal section of pituitaries. Scale bar, (a) 200 μm and (b) 100 μm. (c) Quantification of hormone mRNA levels in 14-day-old Zbtb20+/+ and Zbtb20−/− mice by real-time RT–PCR. Results are presented as fold induction relative to mRNA levels in wild-type mice. A sample was pooled from three to five adenohypophyses of the same phenotype, and the experiment was repeated three times. Values represent mean±s.e.m. *P<0.05, **P<0.01 (Student's t-test). (d) Western blot of hormones and Drd2 proteins in pituitary extracts from 14-day-old Zbtb20+/+ and Zbtb20−/− mice. A sample was pooled from five to eight adenohypophyses of the same phenotype, and the experiment was repeated two times. (e) Anterior pituitary hormonal contents are measured in pituitary extracts by ELISA at P21. n=3. Values represent mean±s.e.m. *P<0.05 versus +/+, **P<0.01 versus +/+ (Student's t-test).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4835541&req=5

f2: Impaired development of endocrine cells in the anterior pituitary of Zbtb20- mice.(a) All five types of cell lineages in anterior pituitary are detected by immunohistochemical staining using GH, PRL, TSHβ, ACTH and LHβ antibodies in 21-day-old Zbtb20+/+ and Zbtb20−/− littermate. PRL-positive cells are absent in mice, but other cell lineages have no apparent changes. (b) Immunohistochemical staining of Drd2 in 21-day-old pituitary sections also shows a loss of positive cells in the mice. Insets show the whole coronal section of pituitaries. Scale bar, (a) 200 μm and (b) 100 μm. (c) Quantification of hormone mRNA levels in 14-day-old Zbtb20+/+ and Zbtb20−/− mice by real-time RT–PCR. Results are presented as fold induction relative to mRNA levels in wild-type mice. A sample was pooled from three to five adenohypophyses of the same phenotype, and the experiment was repeated three times. Values represent mean±s.e.m. *P<0.05, **P<0.01 (Student's t-test). (d) Western blot of hormones and Drd2 proteins in pituitary extracts from 14-day-old Zbtb20+/+ and Zbtb20−/− mice. A sample was pooled from five to eight adenohypophyses of the same phenotype, and the experiment was repeated two times. (e) Anterior pituitary hormonal contents are measured in pituitary extracts by ELISA at P21. n=3. Values represent mean±s.e.m. *P<0.05 versus +/+, **P<0.01 versus +/+ (Student's t-test).

Mentions: To determine the specific deficits in the mutant pituitary, we examined the alteration of its cell composition. Hematoxylin and eosin (H&E) staining revealed a marked decrease in the number of granule-filled acidophilic cells (somatotropes and lactotropes) in the mutant pituitary compared with control at P21 (Supplementary Fig. 2), while basophilic cells (thyrotrophs, adrenocorticotrophs and gonadotrophs) were morphologically normal. We then performed immunohistochemical staining using antibodies specifically against individual hormone produced by each cell type. At P21, there were no apparent differences in the cell densities of GH-, TSH-, LH- or ACTH-immunoreactive cells between mutant and control mice (Fig. 2a). Very strikingly, PRL-immunoreactive cells were completely absent in mutant pituitary at the same age, in contrast to their abundant distribution in normal anterior lobe. The absence of mature lactotropes in mutant pituitary was further confirmed by immunostaining with the antibodies against dopamine receptor D2 (Drd2; Fig. 2b), which is another differentiation marker of lactotropes2829. Consistently, quantitative reverse transcription PCR (RT–PCR) and western blotting analysis revealed that PRL expression was undetectable at either mRNA or protein levels in mutant pituitary at P14 (Fig. 2c,d). In addition, GH expression was decreased by ∼50% in mutant pituitary at both mRNA and protein levels compared to control, POMC mRNA levels increased by 1.46-fold, while LH and FSH mRNA levels were similar between both groups at P14 (Fig. 2c). Furthermore, measurement of the hormone contents in whole pituitary extracts by ELISA revealed that GH contents were decreased by 30% in mutants, PRL contents almost undetected, and TSH, ACTH and LH contents were not altered at P21 (Fig. 2e). These data suggest that ZBTB20 disruption selectively leads to a severe developmental defect of lactotropes, and to some less extent, somatotropes are affected as well.


ZBTB20 is required for anterior pituitary development and lactotrope specification.

Cao D, Ma X, Cai J, Luan J, Liu AJ, Yang R, Cao Y, Zhu X, Zhang H, Chen YX, Shi Y, Shi GX, Zou D, Cao X, Grusby MJ, Xie Z, Zhang WJ - Nat Commun (2016)

Impaired development of endocrine cells in the anterior pituitary of Zbtb20- mice.(a) All five types of cell lineages in anterior pituitary are detected by immunohistochemical staining using GH, PRL, TSHβ, ACTH and LHβ antibodies in 21-day-old Zbtb20+/+ and Zbtb20−/− littermate. PRL-positive cells are absent in  mice, but other cell lineages have no apparent changes. (b) Immunohistochemical staining of Drd2 in 21-day-old pituitary sections also shows a loss of positive cells in the  mice. Insets show the whole coronal section of pituitaries. Scale bar, (a) 200 μm and (b) 100 μm. (c) Quantification of hormone mRNA levels in 14-day-old Zbtb20+/+ and Zbtb20−/− mice by real-time RT–PCR. Results are presented as fold induction relative to mRNA levels in wild-type mice. A sample was pooled from three to five adenohypophyses of the same phenotype, and the experiment was repeated three times. Values represent mean±s.e.m. *P<0.05, **P<0.01 (Student's t-test). (d) Western blot of hormones and Drd2 proteins in pituitary extracts from 14-day-old Zbtb20+/+ and Zbtb20−/− mice. A sample was pooled from five to eight adenohypophyses of the same phenotype, and the experiment was repeated two times. (e) Anterior pituitary hormonal contents are measured in pituitary extracts by ELISA at P21. n=3. Values represent mean±s.e.m. *P<0.05 versus +/+, **P<0.01 versus +/+ (Student's t-test).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4835541&req=5

f2: Impaired development of endocrine cells in the anterior pituitary of Zbtb20- mice.(a) All five types of cell lineages in anterior pituitary are detected by immunohistochemical staining using GH, PRL, TSHβ, ACTH and LHβ antibodies in 21-day-old Zbtb20+/+ and Zbtb20−/− littermate. PRL-positive cells are absent in mice, but other cell lineages have no apparent changes. (b) Immunohistochemical staining of Drd2 in 21-day-old pituitary sections also shows a loss of positive cells in the mice. Insets show the whole coronal section of pituitaries. Scale bar, (a) 200 μm and (b) 100 μm. (c) Quantification of hormone mRNA levels in 14-day-old Zbtb20+/+ and Zbtb20−/− mice by real-time RT–PCR. Results are presented as fold induction relative to mRNA levels in wild-type mice. A sample was pooled from three to five adenohypophyses of the same phenotype, and the experiment was repeated three times. Values represent mean±s.e.m. *P<0.05, **P<0.01 (Student's t-test). (d) Western blot of hormones and Drd2 proteins in pituitary extracts from 14-day-old Zbtb20+/+ and Zbtb20−/− mice. A sample was pooled from five to eight adenohypophyses of the same phenotype, and the experiment was repeated two times. (e) Anterior pituitary hormonal contents are measured in pituitary extracts by ELISA at P21. n=3. Values represent mean±s.e.m. *P<0.05 versus +/+, **P<0.01 versus +/+ (Student's t-test).
Mentions: To determine the specific deficits in the mutant pituitary, we examined the alteration of its cell composition. Hematoxylin and eosin (H&E) staining revealed a marked decrease in the number of granule-filled acidophilic cells (somatotropes and lactotropes) in the mutant pituitary compared with control at P21 (Supplementary Fig. 2), while basophilic cells (thyrotrophs, adrenocorticotrophs and gonadotrophs) were morphologically normal. We then performed immunohistochemical staining using antibodies specifically against individual hormone produced by each cell type. At P21, there were no apparent differences in the cell densities of GH-, TSH-, LH- or ACTH-immunoreactive cells between mutant and control mice (Fig. 2a). Very strikingly, PRL-immunoreactive cells were completely absent in mutant pituitary at the same age, in contrast to their abundant distribution in normal anterior lobe. The absence of mature lactotropes in mutant pituitary was further confirmed by immunostaining with the antibodies against dopamine receptor D2 (Drd2; Fig. 2b), which is another differentiation marker of lactotropes2829. Consistently, quantitative reverse transcription PCR (RT–PCR) and western blotting analysis revealed that PRL expression was undetectable at either mRNA or protein levels in mutant pituitary at P14 (Fig. 2c,d). In addition, GH expression was decreased by ∼50% in mutant pituitary at both mRNA and protein levels compared to control, POMC mRNA levels increased by 1.46-fold, while LH and FSH mRNA levels were similar between both groups at P14 (Fig. 2c). Furthermore, measurement of the hormone contents in whole pituitary extracts by ELISA revealed that GH contents were decreased by 30% in mutants, PRL contents almost undetected, and TSH, ACTH and LH contents were not altered at P21 (Fig. 2e). These data suggest that ZBTB20 disruption selectively leads to a severe developmental defect of lactotropes, and to some less extent, somatotropes are affected as well.

Bottom Line: Disruption of Zbtb20 leads to anterior pituitary hypoplasia, hypopituitary dwarfism and a complete loss of mature lactotropes.Furthermore, endogenous ZBTB20 protein binds to Prl promoter, and its knockdown decreases PRL expression and secretion in a lactotrope cell line MMQ.In addition, ZBTB20 overexpression enhances the transcriptional activity of Prl promoter in vitro.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathophysiology, Second Military Medical University, 800 Xiangyin Road, Shanghai 200433, China.

ABSTRACT
The anterior pituitary harbours five distinct hormone-producing cell types, and their cellular differentiation is a highly regulated and coordinated process. Here we show that ZBTB20 is essential for anterior pituitary development and lactotrope specification in mice. In anterior pituitary, ZBTB20 is highly expressed by all the mature endocrine cell types, and to some less extent by somatolactotropes, the precursors of prolactin (PRL)-producing lactotropes. Disruption of Zbtb20 leads to anterior pituitary hypoplasia, hypopituitary dwarfism and a complete loss of mature lactotropes. In ZBTB20- mice, although lactotrope lineage commitment is normally initiated, somatolactotropes exhibit profound defects in lineage specification and expansion. Furthermore, endogenous ZBTB20 protein binds to Prl promoter, and its knockdown decreases PRL expression and secretion in a lactotrope cell line MMQ. In addition, ZBTB20 overexpression enhances the transcriptional activity of Prl promoter in vitro. In conclusion, our findings point to ZBTB20 as a critical regulator of anterior pituitary development and lactotrope specification.

No MeSH data available.


Related in: MedlinePlus