GRASPing for information: how fat cells sense and respond to nutritional status

Science Spotlight

GRASPing for information: how fat cells sense and respond to nutritional status

From the Rajan Lab, Basic Sciences Division

Oct. 16, 2017

Adipose tissue—also known as fat—serves as the body’s energy storage facility; during times of plenty its inventory is increased, while in times of famine stored fat is broken down to provide energy. In order to perform this function, fat cells need to be able to sense the nutritional status of the organism as a whole. In turn, fat cells communicate the status of the energy stores to other systems in the body by releasing signaling molecules known as adipokines. Despite decades of research, it is not well understood how nutrient status regulates adipokine release at the molecular level. Dr. Akhila Rajan, an Assistant Member in the Basic Sciences Division, started working on this question during her post-doctoral studies with Norbert Perrimon at Harvard Medical School and has continued investigation in her own lab at Fred Hutch. In work recently published in Developmental Cell, Rajan and her colleagues discovered a mechanism by which, in Dr. Rajan’s words, “cells use cytosolic calcium spikes downstream of [the fat breakdown hormone] glucagon as a negative signal for adipokine secretion”.

In humans, the hormone leptin acts on receptors in the brain to inhibit hunger. Mutation of leptin or its receptor is strongly associated with obesity, so understanding the signaling pathways governed by leptin and other adipokines is of great medical interest. Early in her post-doc, Dr. Rajan discovered that in the fruit fly Drosophila, the protein Unpaired 2 (Upd2) performs a similar function as leptin, so she set out to determine how secretion of Upd2 from fat cells is regulated.

microscopy image showing Upd2 localized within GRASP clusters

Image of Drosophila fat tissue showing distribution of adipokine Upd2 (red) in GRASP-containing secretion compartments (green).

Image provided by Dr. Akhila Rajan.

The first step was to determine which secretory route is used to export Upd2 from fat cells. By testing the sensitivity of Upd2 secretion to inhibitors of different pathways, the researchers found that Upd2 bypasses the Golgi and is instead trafficked via a non-traditional secretion route that requires the Golgi reassembly stacking protein (GRASP) (Figure 1). Consistent with a mechanism in which GRASP promotes Upd2 secretion, knockdown of GRASP specifically in Drosophila fat cells led to accumulation of Upd2 inside the cells and lower levels of the fat storage molecule triacylglycerol. Because secretion of Upd2 by fat cells remotely regulates insulin secretion from cells in the fly brain, the authors hypothesized that deletion of GRASP in fat cells, but not in brain cells, should affect insulin accumulation. Indeed, fat cell-specific removal of GRASP led to significantly increased insulin levels.

Having established that GRASP is required for Upd2 to perform its biological functions, the authors asked how GRASP itself is regulated. Using mass spectrometry and biochemistry techniques, they found that GRASP is a target of the calcium-sensing kinase CaMKII. Because CaMKII is activated during starvation, phosphorylation of GRASP was observed to increase under this condition.

To determine the functional consequences of GRASP phosphorylation by CaMKII, the researchers examined Drosophila fat cells under the microscope. They observed that expression of a non-functional form of CaMKII caused fluorescently labeled GRASP to form bright clusters on the apical surface of the fat cells, whereas expression of constitutively active CaMKII decreased cluster formation. These results suggest that only non-phosphorylated GRASP can form apical clusters; indeed, a mutant form of GRASP that mimics phosphorylation was unable to enter the clusters.

Because apical localization of GRASP was more well-pronounced in the well-fed state, when Upd2 is successfully secreted, the authors were able to conclude that the apical clusters represent the functional form of GRASP in terms of promoting Upd2 secretion. During starvation in humans, the catabolic hormone glucagon increases calcium levels and can thus activate CaMKII. Consistent with this notion, the authors observed that knockout of the fly homolog of glucagon, adipokinetic hormone (AKH), causes GRASP to retain apical localization even during starvation, presumably due to reduced CaMKII activity. 

glucagon/AKH inhibits GRASP cluster formation, which is required for leptin/Upd2 secretion and downstream insulin signaling

Overview of the signaling pathway linking nutrient status to adipokine and insulin secretion.

Image provided by Dr. Akhila Rajan.

Importantly, the researchers showed that human leptin expressed in fly cells behaves in the same way as Upd2, suggesting conservation of the discovered mechanism across evolution. Thus, the results of this study have important implications for understanding obesity in humans. According to Dr. Rajan, “previous studies in both mammals and flies have suggested that dysfunctional calcium homeostasis is linked to obesity, Nevertheless, the mechanistic basis for how calcium levels and fat storage are linked has remained unclear. Our study, showing that increased cytosolic calcium, by negatively regulating GRASP via CaMKII mediated phosphorylation, affects Upd2 secretion, provides a specific molecular pathway linking calcium to fat storage” (Figure 2).

In her lab at Fred Hutch, Dr. Rajan is building on these findings with the hope of, in her own words, “understanding how fat communicates nutrient status to the brain, and how chronic nutrient surplus disrupts this communication. Examining this complex question in a simple model system, such as the fruit fly, will provide fundamental insights that can be leveraged to tackle obesity.”

 

Rajan A, Housden BE, Wirtz-Peitz F, Holderbaum L, Perrimon N. “A mechanism coupling systemic energy sensing to adipokine secretion” Developmental Cell. 2017 October 9; 43(1):83-98.e6. doi: 10.1016/j.devcel.2017.09.007.

This research was supported by the National Institutes of Health and the Howard Hughes Medical Institute.