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A new study published in Science Advances has shed light on the complex mechanisms behind the sex differences observed in autism. The research suggests that increased dosage of a gene known as Ube3a can influence autism-related traits in a manner that differs between males and females. By examining mice with extra copies of this gene, researchers found significant sex-specific effects on brain connectivity and behavior, offering insights into why autism is more prevalent in males.
Autism, or autism spectrum disorder (ASD), is a complex neurodevelopmental condition characterized by challenges in social interaction, communication, and a tendency toward repetitive behaviors. These symptoms can vary widely in severity and manifestation, making autism a spectrum disorder.
Despite extensive research, the exact causes of autism remain unclear. It is believed to result from a combination of genetic, environmental, and neurological factors. One intriguing aspect of autism is its higher prevalence in males, with approximately four males diagnosed for every female. This significant sex difference has led scientists to investigate potential genetic and biological mechanisms that might explain this disparity.
One potential genetic mechanism contributing to the male bias in autism is the Ube3a gene. This gene is known to be involved in neurodevelopment and is located on chromosome 15. Variations in the Ube3a gene are associated with several neurodevelopmental disorders, including autism. However, its role in interacting with sex-specific mechanisms had not been thoroughly explored.
“My lab has been studying the genetic and biological causes of autism for many years, especially in relation to the alterations in brain connectivity that characterize these disorders. As part of this research, we have been scanning the brain of mice harboring a duplication in autism-risk gene Ube3a,” said study author Alessandro Gozzi, a senior scientist and director of the Functional Neuroimaging Laboratory at the Italian Institute of Technology.
“These investigations led us to serendipitously discover significant effect of sex on the patterns of alterations we mapped, with male and female mice exhibiting different and diverging effects. This finding intrigued us and prompted us to further explore the functions of Ube3A. We then realized that this gene plays a poorly investigated role in controlling the expression of other genes in a sex-dependent manner. Through further investigations, we found that Ube3A acts as a sex-specific switch that controls the expression of many other genes important for autism in both rodents and humans.”
To investigate the effects of increased dosage of the Ube3a gene, the researchers used genetically modified mouse model to mimic the human condition of 15q duplication syndrome, which includes duplications of the Ube3a gene and is associated with autism. Specifically, the researchers used Ube3a2X mice, which carry two extra copies of the Ube3a gene, comparing them to a sample of wild-type controls.
Functional magnetic resonance imaging (fMRI) was used to study brain connectivity. Both male and female mice underwent resting-state fMRI (rsfMRI) to measure the functional connections between different brain regions. Structural MRI was also performed to examine any anatomical changes in the brain.
The fMRI results revealed significant differences in brain connectivity between male and female Ube3a2X mice. Female mice with increased Ube3a dosage exhibited reduced connectivity in key brain regions, including the hypothalamus and prefrontal cortex. In contrast, male mice showed increased connectivity in these areas. These findings suggest that the Ube3a gene dosage affects brain connectivity differently in males and females, which could contribute to the sex-specific prevalence of autism.
“The highly divergent effect of the same mutation on brain function in rodents was really striking and surprising, and it became a major lead for our subsequent investigations,” Gozzi told PsyPost.
To assess autism-related behaviors, the researchers conducted a series of behavioral tests. These included the open-field test to measure locomotion and anxiety, the rotarod test to evaluate motor coordination, and the self-grooming scoring to observe repetitive behaviors. Additionally, social behaviors were assessed using the habituation/dishabituation social interaction test and the three-chamber social interaction test.
Male Ube3a2X mice displayed significantly more repetitive behaviors, such as excessive grooming, compared to their wild-type counterparts. This increase in stereotyped behavior was not observed in female Ube3a2X mice. The rotarod test also revealed motor coordination impairments in Ube3a2X mice, but these were not sex-specific. In terms of social behavior, both the habituation/dishabituation test and the three-chamber test showed no significant differences in social interactions between Ube3a2X mice and wild-type controls, regardless of sex.
The researchers also performed RNA sequencing to analyze gene expression in specific brain regions, particularly the prefrontal cortex and hypothalamus. This allowed them to identify any differences in gene activity between the Ube3a2X mice and their WT counterparts, as well as between males and females.
The gene expression analysis provided further insights into the sex-specific effects of increased Ube3a dosage. In the prefrontal cortex, many autism-associated genes exhibited dysregulated expression in a sex-dependent manner. For instance, some genes were upregulated in male Ube3a2X mice but downregulated in females, and vice versa. This differential gene expression was particularly evident in genes located on the X chromosome and those influenced by sex steroid hormones, such as androgen and estrogen receptors.
“The main takeaway is that the sex bias in autism (with boys being four times more affected than girls) can plausibly be explained by genetic mechanisms, specifically those interacting with sex hormones,” Gozzi explained. “This finding adds to a growing body of evidence suggesting that autism is, in many respects, a genetic condition.”
The findings advance our understanding of the genetic and biological factors underlying the sex bias in autism. However, Gozzi noted that “this is just one of possibly many genetic mechanisms leading to sex bias in autism. The challenge now is to understand which and how many other genetic and biological mechanisms contribute to this phenomenon.”
“Our long-term goal is to better understand how genetic alterations that confer risk of developing autism alter brain function. This remains a major knowledge gap that we need to address.”
The study, “Sex-biasing influence of autism-associated Ube3a gene overdosage at connectomic, behavioral, and transcriptomic levels,” was authored by Caterina Montani, Luigi Balasco, Marco Pagani, Filomena Grazia Alvino, Noemi Barsotti, A. Elizabeth de Guzman, Alberto Galbusera, Alessia de Felice, Thomas K. Nickl-Jockschat, Sara Migliarini, Simona Casarosa, Pierre Lau, Lorenzo Mattioni, Massimo Pasqualetti, Giovanni Provenzano, Yuri Bozzi, Michael V. Lombardo, and Alessandro Gozzi.
