Skip to main page content
U.S. flag
See More

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2008 Oct 21;105(42):16314-9.
doi: 10.1073/pnas.0808517105. Epub 2008 Oct 13.

PERK-dependent regulation of lipogenesis during mouse mammary gland development and adipocyte differentiation

Ekaterina Bobrovnikova-Marjon  1 , Georgia Hatzivassiliou, Christina Grigoriadou, Margarita Romero, Douglas R Cavener, Craig B Thompson, J Alan Diehl
Affiliations

PERK-dependent regulation of lipogenesis during mouse mammary gland development and adipocyte differentiation

Ekaterina Bobrovnikova-Marjon et al. Proc Natl Acad Sci U S A. .

Abstract

The role of the endoplasmic reticulum stress-regulated kinase, PERK, in mammary gland function was assessed through generation of a targeted deletion in mammary epithelium. Characterization revealed that PERK is required for functional maturation of milk-secreting mammary epithelial cells. PERK-dependent signaling contributes to lipogenic differentiation in mammary epithelium, and perk deletion inhibits the sustained expression of lipogenic enzymes FAS, ACL, and SCD1. As a result, mammary tissue has reduced lipid content and the milk produced has altered lipid composition, resulting in attenuated pup growth. Consistent with PERK-dependent regulation of the lipogenic pathway, loss of PERK inhibits expression of FAS, ACL, and SCD1 in immortalized murine embryonic fibroblasts when cultured under conditions favoring adipocyte differentiation. These findings implicate PERK as a physiologically relevant regulator of the lipogenic pathway.

PubMed Disclaimer

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Conditional deletion of perk in mouse mammary gland affects mouse milk fat composition, mammary gland lipid content, and pup growth. (A) Whole mounts of the mammary glands from control (Flox Cre−) and cKO (Flox Cre+) mice on lactation day 3. (B) H&E staining of sections from control and cKO glands on lactation day 3. Arrows indicate the areas of uncollapsed adipocyte stroma. (C and D) Free fatty acids or triglyceride content of milk collected from 4-month-old cKO (Flox Cre+, n = 3) and control (Flox Cre−, n = 5) dams on lactation day 12. Error bars represent SD. (E) Nile red staining of mammary glands from 4-month-old control and cKO dams on lactation day 12. Lu, lumen of the alveoli. (F) Average litter weight plotted vs. pups' age in a cohort of 4-month-old control (Cre−, n = 8) dams nursing wild-type pups and cKO (Cre+, n = 9) animals nursing cKO pups. Error bars represent SD.
Fig. 2.
Fig. 2.
Perk deletion in mouse mammary epithelium inhibits SREBP1 activation and the sustained induction of lipogenic enzymes. (A) RT-PCR analysis for SREBP1, FAS, ACL, SCD1, and XOR from mammary extracts of 4-month-old control (Flox Cre−, n = 3) and cKO (Flox Cre+, n = 3) dams at P16, lactation day 3 (L3), and 12 (L12). Error bars represent SEM (*, P < 0.005; #, P < 0.05). (B) Western blot for PERK, FAS, ACL, SCD1, XOR, and eIF4E of mammary gland extracts from the same developmental stages as in A. IP-Western for SREBP1 was performed on the same samples. NRS, nonspecific rabbit serum. (C) Western blot for PERK, Myc-tagged SREBP1 precursor (SREBP1128) or processed isoform (SREBP168), phosho-eIF2α, and eIF4E on samples from perk LoxP pBabe MEFs (Mock) or perk LoxP Cre MEFs (Cre) expressing Myc-SREBP1 and treated as indicated with 50 nM thapsigargin (Th) and 10 μM MG132. (D) Western blot for Myc-SREBP1 precursor and processed isoform, phosho-eIF2α, and eIF4E on samples from eIF2α wild-type (wt) or eIF2α S51A mutant knockin MEFs expressing Myc-SREBP1 and Myc-Insig1. Cells were treated with 50 nM thapsigargin (Th) and 10 μM MG132 where indicated.
Fig. 3.
Fig. 3.
PERK-dependent depletion of Insig1 contributes to induction of SREBP1 target genes. (A) Western blot for proteins indicated on samples from wild type (wt) or perk knockout (KO) MEFs treated as indicated. (B) RT-PCR analysis of FAS, ACL, and SCD1 in perk LoxP pBabe MEFs (Mock), perk LoxP Cre MEFs (Cre), or perk/Gcn2 double-knockout (DKO) MEFs. Error bars represent SD for three independent experiments. (C) EMSA analysis of nuclear extracts isolated from perk LoxP pBabe MEFs or perk LoxP Cre MEFs treated with thapsigargin (Th). + Cold indicates the reaction performed in the presence of unlabeled IRS2 duplex; + α-SREBP1 indicates the addition of SREBP1 antibody.
Fig. 4.
Fig. 4.
PERK affects the lipogenic program during differentiation of immortalized MEFs into adipocytes. (A) Western blot for FAS, ACL, Myc-SREBP1 precursor, PERK, phosho-eIF2α, total eIF2α, and eIF4E in perk LoxP pBabe MEFs (Mock) or perk LoxP Cre MEFs (Cre) infected with virus encoding Myc-SREBP1 and cultured in adipocyte differentiation mixture for 0, 3, 5, 7, 9, 11, or 13 days. (B) RT-PCR quantification of FAS, ACL, SCD1, and SREBP1c from cells treated as in A. (C) Oil Red O staining on perk LoxP pBabe MEFs or perk LoxP Cre MEFs expressing Myc-SREBP1 and cultured in adipocyte differentiation mixture.
See this image and copyright information in PMC

References

    1. Ma Y, Hendershot LM. The unfolding tale of the unfolded protein response. Cell. 2001;107:827–830. - PubMed
    1. Schroder M, Kaufman RJ. The mammalian unfolded protein response. Annu Rev Biochem. 2005;74:739–789. - PubMed
    1. Wang Y, et al. Activation of ATF6 and an ATF6 DNA binding site by the endoplasmic reticulum stress response. J Biol Chem. 2000;275:27013–27020. - PubMed
    1. Wu J, et al. ATF6alpha optimizes long-term endoplasmic reticulum function to protect cells from chronic stress. Dev Cell. 2007;13:351–364. - PubMed
    1. Harding HP, Zhang Y, Ron D. Protein translation and folding are coupled by an endoplasmic-reticulum-resident kinase. Nature. 1999;397:271–274. - PubMed

Publication types

MeSH terms