Building the case against a rogue gene

Three labs, including one at Yale, independently target a gene implicated in autism.

Researchers know that defects in brain development are to blame for autism, but pinpointing the likely genetic culprits has remained an elusive goal. To put it in mobster terms, there is no single “Tony Soprano gene” corrupting brain development. Instead, scientists believe that multiple “small-time thug genes” gang up to undermine the developing brain. Because the individual effects of these crooked genes may be subtle, it has been hard to get the goods on them.

To help crack the case, Matthew W. State, M.D., Ph.D. ’01, the Irving B. Harris Associate Professor of Child Psychiatry in the Child Study Center and of genetics and director of the Program on Neurogenetics, sought to flush out autism-associated genes by focusing on clues from certain affected individuals. Children with autism—or any of a spectrum of related disorders—have difficulty communicating and interacting with others, exhibit stereotyped behaviors and often suffer from mental retardation and seizures. A small percentage of these children also have a visible chromosomal abnormality. In one such patient State found that the abnormality disrupted the Contactin Associated Protein-Like 2 (CNTNAP2) gene, which encodes a protein that helps brain signals pass from one neuron to another. Based on prior work by himself and others linking contactin proteins to autism spectrum disorders, mental retardation and seizures, State grew suspicious of CNTNAP2.

With the help of colleagues in clinical medicine, neurobiology, biochemistry and genetics, State has collected a body of evidence that strongly incriminates CNTNAP2 as one of perhaps many autism accomplices. First, CNTNAP2 is present at the scene of the crime, including all layers of the cerebral cortex within the temporal lobe and within the limbic system, a brain circuit involved in social behavior. Second, CNTNAP2 is found with its binding partner, contactin 2, at synaptic plasma membranes—the gates of communication between neurons. The third and strongest line of evidence against CNTNAP2 is that sequencing of the gene from 635 autistic patients and 942 controls turned up 13 rare, unique changes to the encoded protein that were found only in autistic individuals. Eight of these mutations are predicted to disrupt the proper functioning of CNTNAP2. One particular mutation was identified in four autistic children in three unrelated families, but not in more than 4,000 chromosomes from controls. “This is strong but not definitive evidence linking this gene with autism,” according to State.

Unbeknownst to each other and to State, two other medical research laboratories—the labs of Daniel H. Geschwind, M.D., Ph.D., at the University of California, Los Angeles (UCLA), and of Aravinda Chakravarti, Ph.D., at Johns Hopkins University—also fingered CNTNAP2 as causative of autism. Both Geschwind and Chakravarti independently homed in on CNTNAP2 after surveying the genomes of hundreds of individuals and identifying a particular chunk of genetic material that appeared to surface in families with autism. State and Geschwind, longtime friends who met as residents at UCLA, learned of the other’s discovery while catching up during one of their regular phone conversations. Soon after, Geschwind caught wind of Chakravarti’s work through the research grapevine. When the three scientists compared notes, they decided to co-publish their findings to build the strongest case possible against CNTNAP2. “There’s a reason we’re all landing on this gene,” said State. All three papers were published in the January issue of the American Journal of Human Genetics.

State thinks that identification of CNTNAP2 may give him the traction he needs to begin to understand the complex biology of autism. “Our hope is that our continued work on understanding the biology of CNTNAP2 will lead to real opportunities for novel approaches to treatment,” said State.

—Kara A. Nyberg

In the olfactory bulb, new neural stem cells learn to listen before they speak

Like a newborn learning from its parents, a neuron born of neural stem cells in the adult brain must take its cues from its elders if it hopes to mature and survive, according to new research headed by Charles A. Greer, Ph.D., professor of neurosurgery and neurobiology.

In findings published in the September 12, 2007, issue of The Journal of Neuroscience, Greer and Mary C. Whitman, an M.D./Ph.D. candidate in his lab, tracked the development of new neurons in a region of the brain called the olfactory bulb, which receives information about odors from the nose. It is one of the few regions in the adult brain that allows new neurons to be generated and integrated into existing neural circuits.

However, such assimilation is not easy. New brain cells destined for the olfactory bulb have to migrate vast distances from their birthplace, and half of these newborn neurons die between 15 and 45 days after being generated, presumably because they fail to integrate within the neural circuitry.

Greer and Whitman found that although the long, spindly arms of new neurons are present 10 days after being generated, they don’t form the connections that let them talk to other olfactory bulb neurons until three weeks after birth. Even then, it takes six to eight weeks for the cells to mature and achieve complete integration.

“New neurons are essentially taught to listen before they’re allowed to talk,” said Greer. He and Whitman found that fibers extending from older neurons located in higher centers of the brain first connect to short arms projecting from the base of new neurons about 10 days after generation. Whitman and Greer believe that these early synapses provide a conduit through which elder brain cells control the development, and ultimately the survival, of new neurons within existing brain circuitry. This ensures that the new lines of communication don’t garble pre-existing lines.

According to Greer, these findings have important implications for using adult neural stem cells to replace brain cells lost by trauma or neurodegeneration, such as in Parkinson’s disease. “To use stem cells in a transplant strategy, we’re going to have to understand the kinds of synapses new brain cells make as well as the kinds of synapses they receive from existing circuits. Our goal is to prevent these cells from being potentially disruptive by getting into the wrong synaptic circuit or by acting in a precocious way,” he said.

—K.A.N.

et cetera

Big role for tiny RNA

Tiny RNAs discovered in “junk” DNA play an important role in controlling gene function, Yale scientists reported in the journal Nature in October.

A team led by Haifan Lin, Ph.D., director of the Yale Stem Cell Center and professor of cell biology, discovered these RNAs, called piRNAs, in mammalian reproductive cells in 2006. The team’s findings suggest that piRNAs also exist in nonreproductive body cells and help to control stem cell fate and tissue development. The researchers found that a particular piRNA forms a complex with a protein called Piwi, which then binds to a specific region of chromatin (i.e., the genome) that regulates gene activity.

“This finding revealed a surprisingly important role for piRNAs, as well as junk DNA, in stem cell division,” Lin said. “It calls upon biologists to look for answers beyond the 1 percent of the genome with protein-coding capacity to the vast land of junk DNA, which constitutes 99 percent of the genome.”

—John Curtis

Virus kills brain tumors

Yale researchers have engineered a virus that can find its way through the vascular system and kill deadly brain tumors, offering a potential new treatment for cancers in the brain.

Such malignant brain tumors as glioblastomas and metastatic tumors are diagnosed in 22,000 Americans each year. There is no cure for these malignancies. They often kill within months; current treatments usually fail because they don’t kill all the cancer cells.

Anthony N. van den Pol, Ph.D., professor of neurosurgery, and colleagues reported in the Journal of Neuroscience in February that they had transplanted multiple types of human and mouse tumors into the brains of mice and then inoculated the mice with a lab-created vesicular stomatitis virus known as vsvrp30a, a distant cousin of the rabies virus. Three days later, the tumors had been infected by the virus and “were dying or dead,” while transplanted normal cells were spared, van den Pol said. “This underlines the virus’ potential therapeutic value against multiple types of brain cancers.”

—John Dillon


			Originally published in Yale Medicine, Spring 2008. Copyright © 2008 Yale University School of Medicine. All rights reserved.