Despite having identical genetic instructions, female honey bee larvae can develop into either long-lived reproductive queens or short-lived sterile workers who help rear their sisters rather than laying their own eggs. Now, an interdisciplinary team led by researchers at Penn State has uncovered the molecular mechanisms that control how the conflict between genes inherited from the father and the mother determine the larva's fate.
They published their findings this week (June 18) in Genome Biology.
"Imagine if your mother's genes and your father's genes were in constant disagreement about how you should develop - that is essentially what genomic imprinting is, and we see that it happens across the tree of life: from honey bees to humans," said Sean Bresnahan, the lead author of the study who conducted the study as a doctoral candidate in the Interdisciplinary Graduate Degree Program in Molecular, Cellular, and Integrative Biosciences in the Huck Institutes of the Life Sciences at Penn State. Supported by a U.S. National Science Foundation (NSF) Graduate Research Fellowship at the time, Bresnahan graduated in 2024 and is now a data scientist at the University of Texas MD Anderson Cancer Center. "We found that this genetic 'argument' can be detected during a critical developmental window where a honey bee larva becomes either a queen or a worker."
That critical window closes and the bee's fate becomes irreversible 192 hours after the egg is laid. To distinguish between patrigenes - genes inherited from the father - and matrigenes - genes inherited from the mother, study co-author Kate Anton, a research technologist in the Center for Pollinator Research at Penn State, used instrumental insemination to create specific genetic crosses between selected queens and male bees, known as drones. The researchers worked with the Penn State Genome Research Incubator to analyze the larvae's RNA, which contains and uses inherited genetic information to create proteins and support cellular activity, and identify the genes that were expressed differently between the two groups. The researchers also sequenced the parents' genomes and used genetic markers to trace parent-of-origin gene expression in the larvae, meaning they could see how gene expression differed depending on whether the gene was from the mother or father.
"We found patrigenes were expressed at higher levels in queen-destined larvae, and matrigenes were expressed at higher levels in worker-destined larvae," Bresnahan said.
The researchers then examined cellular and physiological pathways to determine if the genes showing parent-of-origin expression were functioning in the same pathway.
"We saw a striking match between the expression of matrigenes and patrigenes in the same pathway," Bresnahan said, explaining that if a matrigene had increased expression then a patrigene would have decreased expression in the same pathway, or vice versa, showing that the two genes were working against each other. "If one gene showed parent-specific expression, another gene in the same pathway showed parent-specific expression from the opposite parent."
Previously, the team examined whether DNA methylation - a process in which protein tags change how a gene is expressed without altering the underlying DNA - was the molecular mechanism underlying these differential gene expression patterns.
"In mammals and plants, differential expression in imprinted genes - genes where only the information inherited from one parent is expressed - is typically mediated by differences in DNA methylation in the regulatory regions, where specific DNA sequences control expression, of these genes," said study co-author, Christina Grozinger, Publius Vergilius Maro Professor of Entomology and director of the Huck Institutes of the Life Sciences. "But our previous work found that DNA methylation does not have this function in honey bees."
The researchers then turned to the idea that it may not be tags on the DNA that influence gene expression in honey bees, but rather tags on the structures around which DNA packages itself. DNA is wound around histone proteins, in a structure called chromatin. During cellular reproduction, chromatin condenses into chromosomes. Typically developing multicellular organisms inherit equal numbers of chromosomes from each parent, providing the pool of genes from which an organism's specific genetic composition is made up. The team hypothesized that changes in the structure of histone proteins - due to chemical "tags" placed on the tails of the proteins as the result of cellular processes - could make the patrigenes and matrigenes more or less accessible to other regulatory factors. Such control could potentially change their expression.
To test this hypothesis, the team used a method developed by study co-author, Shaun Mahony, associate professor in the Center for Eukaryotic Gene Regulation and the Department of Biochemistry and Molecular Biology at Penn State. The process involves fragmenting DNA that is bound to a protein - histone proteins, in this case - and then using antibodies that are specific to the tagged proteins to capture and isolate these from the rest of the chromatin. Researchers can then analyze the DNA crosslinked to the captured protein and determine which genes are involved and whether they are expressed or inhibited.
"We found that, in honey bees, parent-of-origin expression is regulated by histone modifications," Bresnahan said.
He explained that the chemical tags modifying the histone proteins appears to mediate whether patrigenes or matrigenes are expressed and, ultimately, whether a honey bee becomes a queen or a worker. While the underpinning mechanism is not DNA methylation like the researchers initially thought, Bresnahan said it's not a surprising finding.
