Two brain proteins associated with distinct neural functions have been found to interact physically and functionally. While both can regulate nerve cell firing and are implicated in learning, one, a structural protein for the excitatory neurotransmitter glutamate, reduced the function of the other, a protein receptor for dopamine. The findings, published in the May 25 Journal of Biological Chemistry, reveal that the structural protein, PSD-95, may be an effective drug target for behavioral disorders associated with dopamine imbalances in the brain.
Though it has been known that the dendritic side of glutamate synapses contain copious dopamine receptors with profound modulatory effects, how the dopamine receptors are regulated within these synapses has remained unclear. Previously, Wei-Dong Yao, HMS assistant professor of psychiatry at the New England Primate Research Center (NEPRC), found that PSD-95 in glutamatergic synapses was involved in dopamine signaling. “Knockout mice without PSD-95 were more sensitive to psychostimulants that increase dopamine signaling than normal mice,” said Yao, who has shown PSD-95’s role in dopamine signaling through utilization of cocaine and amphetamines.
Now Yao, postdoctoral fellow Jingping Zhang, and colleagues report that D1 dopamine receptors and PSD-95 coprecipitated in vivo in such areas as the dopamine-rich striatum and cortex and in vitro in cell lines. Looking for a more detailed view of the proteins’ entanglement, the researchers found that the molecules linked together at PSD-95’s N-terminus and at the intracellular C-terminal tail of the D1 receptor.
“Here’s the novel thing,” Yao explained. “The interaction did not involve any of the well-characterized domains. Instead, it involved a novel protein domain of PSD-95.” The functional implications of the N-terminus domain are not known, but some hints of function are provided by the site of interaction on the D1 receptor.
The C-terminal tail of the D1 receptor is known for trafficking dopamine receptors to and from active duty at the cell membrane. When radioligand studies showed no dopamine-binding differences or differences in the total number of dopamine receptors with and without PSD-95 expression, Yao and his research team wondered whether the availability of active receptors at the cell surface explained the differences in dopamine signaling. Indeed, the scientists found through biochemical assays and confocal microscopy that PSD-95 lowered the levels of D1 receptors on the cell membrane.
The identification of PSD-95 as a molecule that balances two important neurotransmitter systems could have implications for a variety of psychiatric disorders, including schizophrenia and addiction. “In addition to drugs which act primarily as dopamine receptor agonists or antagonists,” said Gregory Miller, HMS assistant professor of psychiatry at NEPRC and a co-author on the paper, “PSD-95 now represents a new potential target for medications that modulate dopaminergic function.”
Heterochromatin has puzzled scientists for decades. It was identified almost 90 years ago by its darker appearance under the microscope in relation to euchromatin. Only later did scientists begin to understand that this tightly packaged DNA silences gene expression and has roles in chromosome maintenance. Today, heterochromatin continues to maintain its silence, only reluctantly giving up secrets to its quiescence.
Danesh Moazed, HMS professor of cell biology, and colleagues have just uncovered two mechanisms that contribute to heterochromatin silencing in fission yeast. Paradoxically, both depend on the very thing heterochromatin is thought to prevent, transcription.
The first mechanism relies on RNA interference. Though RNAi has been linked to heterochromatin silencing before—key RNAi enzymes, including Dicer and Argonaute, are essential for heterochromatin formation—it is unclear whether RNAi targets heterochromatic transcripts themselves or merely regulates other genes that contribute to heterochromatin formation. If the former, then cells would have to make small interfering RNAs (siRNAs) that complement heterochromatin genes; however, various research labs, including Moazed’s, have failed to detect such siRNAs in yeast—until now.
To pinpoint heterochromatin siRNAs, Moazed and colleagues, including HMS associate professor of cell biology Steven Gygi, turned to a more sensitive detection method and to yeast cells that are deficient in exoribonuclease, which degrades siRNA. Writing in the May 18 Cell, postdoctoral research fellow Marc Buhler and colleagues say that when they inserted the reporter gene ura4+ into yeast centromeric heterochromatin, they were able to detect ura4+ siRNAs with the more sensitive assay, albeit in low amounts. Nevertheless, “because siRNAs are a signature of processing by RNAi, this establishes RNAi as one part of the mechanism that contributes to heterochromatin silencing,” said Moazed.
But it turns out that this is only part of the picture, because when Buhler inserted the ura4+ transgene into a different heterochromatin location called the silent mating–type locus, ura4+ siRNAs were not detected. And that’s where the second mechanism comes in. The researchers wondered if the exosome, a nuclear ribonuclease complex that degrades aberrant RNA transcripts, might also play a role in gene silencing. Specifically, they tested whether a protein called Cid14 might be involved because it is a homolog of a budding yeast polyadenylase that marks transcripts for exosome-mediated degradation. Sure enough, by knocking out Cid14 in fission yeast, the researchers abolished silencing of the ura4+ reporter.
If these mechanisms are conserved in humans, it could have important implications for RNAi-based therapies. “It suggests that you may be able to design RNAi therapies that can have permanent effects at the chromatin level, leading to very stable changes in gene expression patterns,” said Moazed.
One warning sign of impending type 2 diabetes is the accumulation of fat in muscle tissue. That flab is more than just excess baggage, according to new work from Joslin Diabetes Center researchers. In a paper in the May 25 Journal of Biological Chemistry, HMS assistant professor of medicine Mary-Elizabeth Patti and colleagues show that fats, and specifically saturated fats, weigh down a key gene regulator in muscle cells. Their results show how a high-fat diet may trigger the metabolic changes seen in obesity and type 2 diabetes.
Patti and colleagues had previously found that levels of two gene activators, PGC-1 alpha and beta, were reduced in people at risk for type 2 diabetes even before they developed the disease. PGC-1 proteins regulate an array of metabolic genes responsible for proper mitochondrial function, and their levels have a major impact on energy balance. Both genes and environment affect PGC-1 expression, and thus the risk of diabetes.
In the environmental corner, obesity, overeating, and a high-fat diet all diminish PGC-1–gene expression. In their new study, Patti and coworkers searched for the specific nutrients responsible for cutting PGC-1 in muscle cells. They ruled out sugar, insulin, and excess amino acids before finding that high levels of saturated fats depressed transcription of the PGC-1 genes.
“The saturated fatty acids, also known as the ‘bad’ fats, reproduce the same pattern of gene expression in our muscle cells as we see in obesity, in people on high-fat diets, and in type 2 diabetes,” Patti said.
The net result is impaired mitochondrial function and a decreased fat-burning capacity, which could exacerbate the build-up of fat in muscle cells.
The good news is that the effects of saturated fats on PGC-1 can be reversed by giving the cells good fats, such as polyunsaturated omega-3s, by exercise, and by drugs commonly used to treat type 2 diabetes.