Mediobasal hypothalamic FKBP51 acts as a molecular switch linking autophagy to whole-body metabolism

Autophagy and its link to disease

Autophagy is an important cellular pathway that is activated in response to a variety of stressors, most notably starvation and nutrient deficiency. Under these conditions, autophagy is used as a recycling process to degrade unnecessary and damaged material that can be reused for more important cellular processes like energy metabolism. Impaired or disrupted autophagy is associated with disease states including cancer, neurodegeneration, cardiac diseases, and metabolic syndrome.

Autophagy is linked to metabolic syndrome

Autophagy plays an important role in adipocyte differentiation indicating it might be a key player in obesity and metabolic syndrome. These conditions are regulated by complex neuronal circuits integrated in the hypothalamus to maintain energy homeostasis. The two main neurons that comprise these pathways are agouti-related peptide (AgRP) neurons that are activated by negative energy balance states like fasting and pro-opiomelanocortin (POMC) neurons that are activated by positive energy states.¹

Genetic studies have shown variable effects of ATG7 depletion depending on tissue and cell type. ATG7 depletion in adipose tissue resulted in resistance to diet-induced obesity and elevated levels of brown adipose tissue. ATG7 depletion in skeletal muscle resulted in a similar phenotype and these mice were also resistant to diet-induced obesity and insulin resistance. Conversely, ATG7 depletion in the hypothalamus resulted in higher fat mass and body weight.² Additionally, ATG7 deficiency in the mediobasal hypothalamus (MBH) had differing effects depending on the neuronal type. Obesity was induced in ATG7-/- POMC neurons, but in ATG7-/- AgRP neurons animals had decreased body weight.³ These data suggest that autophagy plays a key role in obesity and metabolic syndrome, but the precise mechanisms and regulatory proteins are still being elucidated.

FKBP51 (FK506-binding protein 51) is a regulator of obesity and glucose metabolism. It has also been found to induce autophagy by altering inhibitory and degrative phosphorylation of Beclin-1. Since FKBP51 and autophagy are upregulated after starvation in the MBH, the main region for metabolic control in the brain, the authors investigated the role FKBP51 plays in autophagy and the link to obesity.

Linking FKBP51 to known molecular regulators of autophagy under metabolic stress

TSC2 complex

Under glucose starvation, AMP-activated protein kinase (AMPK) and liver kinase B1 (LKB1) sense the ATP/AMP ratio and activate the TSC2 (tuberous sclerosis 2) complex. The TSC2 complex inhibits mTOR (mechanistic target of rapamycin) thereby inducing autophagy.⁴ In the current study, under nutrient deprivation, FKBP51 knockout cells had decreased LKB1 and AMPK activation and reduced TFEB (transcription factor EB) nuclear translocation. This indicates decreased autophagy signaling. Conversely, FKBP51 overexpressing cells had increased pro-autophagic phosphorylation of Beclin-1, along with decreased mTOR activity, increased ATG16L1 phosphorylation and decreased levels of the autophagy substrate p62. These molecular signatures point towards increased autophagic activity. Together, these data suggest that FKBP51 regulates autophagy in response to metabolic challenge.

WIPI proteins

WIPI proteins (tryptophan-aspartic acid (WD)-repeat proteins that interact with phosphoinositide protein family) are key adaptor proteins that connect autophagy proteins with proteins that are sensitive to metabolic changes. The interaction of FKBP51 with WIPI3 and WIPI4 provides a mechanism for how FKBP51 regulates autophagy and whole-body metabolism. Immunoprecipitation experiments showed FKBP51 interacts with WIPI3 and WIPI4, but not WIPI1 or WIPI2 (which have known roles in autophagy); with AMPKalpha1, AMPKbeta1, and AMPKgamma2; and with LKB1. AMPK-FKBP51 interactions were reduced in WIPI4 knockdown cells, but the FKBP51-LKB1 interaction remained. The AMPK-WIKI4 interaction is dependent on FKBP51 suggesting that FKBP51 creates a scaffold linking the WIKI network and AMPK signaling. WIPI3 drives the interaction between FKBP51 and TSC2, but WIPI3 can interact with TSC1 and TSC2 in the absence of FKBP51.

Regulation of autophagy in the mediobasal hypothalamus by FKBP51

To assess the effects of diet on autophagy and FKBP51 levels, C57BL/6N mice were fed a high fat diet (58% calories from fat) and MBH regions were examined. These mice had increased FKBP51 levels and decreased p62 levels in the MBH at 10 weeks. FKPB51 was then depleted specifically from the MBH by injecting a Cre-expressing virus into the MBH of FKBP51^lox/lox mice. FKBP51 depletion in the MBH resulted in substantial weight gain on a normal chow diet. This was unexpected because full body-deficient FKBP51 mice had a lean body type when fed a high fat diet. Additionally, mice overexpressing FKPB51 in the MBH and fed a high fat diet had decreased body weight and improved glucose tolerance compared to wild type mice. These data show that there are tissue-specific effects of FKBP51 depletion and overexpression and that FKBP51 in the MBH is important for dealing with obesogenic stressors.

To further assess the connection between metabolic stress and autophagy, the mice depleted of FKBP51 in only the MBH (FKBP51 MBH-deficient mice) were used to examine the regulation of autophagy signaling. FKBP51 MBH-deficient mice had decreased LKB1 and AMPK binding to WIPI4 in the MBH, ultimately resulting in decreased pro-autophagic phosphorylation of ULK1, Beclin-1, and TSC2. These FKBP51 MBH-deficient mice also had decreased LC3B-II levels and increased p62 levels. Interestingly, FKBP51 overexpressing mice also had decreased LKB1 and AMPK binding to WIPI4 and increased p62 levels. However, LC3B-II levels were increased, and chloroquine studies showed defective autophagosome-lysosome fusion in the FKBP51 overexpressing mice. Therefore, mice lacking FKBP51 or overexpressing FKBP51 in the MBH had impaired autophagy. The authors hypothesized that the amount of FKBP51 overexpression had a direct correlation with the amount of autophagy signaling. Titration experiments found that moderate (~2-fold) increase in FKBP51 levels induced autophagy but 3-4-fold increased levels of FKBP51 inhibited autophagy.

This study reveals tissue-specific autophagy signaling pathways in response to obesity and that FKBP51 is a regulatory link between stress-induced LKB1/AMPK mediated autophagy induction and WIPI protein scaffolds.

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References

1. Caron A, Jane Michael N. New Horizons: Is Obesity a Disorder of Neurotransmission? J Clin Endocrinol Metab. Published online June 12, 2021. doi:10.1210/clinem/dgab421
2. Menikdiwela KR, Ramalingam L, Rasha F, et al. Autophagy in metabolic syndrome: breaking the wheel by targeting the renin–angiotensin system. Cell Death Dis. 2020;11(2):87. doi:10.1038/s41419-020-2275-9
3. Häusl AS, Bajaj T, Brix LM, et al. Mediobasal hypothalamic FKBP51 acts as a molecular switch linking autophagy to whole-body metabolism. Sci Adv. 2022;8(10). doi:10.1126/sciadv.abi4797
4. Dikic I, Elazar Z. Mechanism and medical implications of mammalian autophagy. Nat Rev Mol Cell Biol. 2018;19(6):349-364. doi:10.1038/s41580-018-0003-4