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  • Examining the Mechanism of Spore Sacs Undergoing Sk-3-Based Spore Killing After Deletion Of Neurospora crassa DNA Intervals i382 and i400
Title: Examining the Mechanism of Spore Sacs Undergoing Sk-3-Based Spore Killing After Deletion Of Neurospora crassa DNA Intervals i382 and i400
Neurospora crassa is a well-known model organism for studying eukaryotic genetics, particularly non-Mendelian inheritance mechanisms such as meiotic drive. In N. crassa, meiotic drive can be observed in fungal spore killing, where Spore killer-3 (Sk-3) is a selfish genetic element transmitted to offspring through spore killing. Sk-3 is thought to contain two principal components: a killer (poison) gene and a resistance (antidote) gene. While the resistance gene (rsk) has been identified, the killer gene remains unknown. Building on previous research that identified a 1.3 kb DNA interval (i350) essential for Sk-3-based spore killing, I analyzed two subintervals, i382 and i400, to narrow down the functional components of the Sk-3 locus. Deletion of i382 does not disrupt spore killing and deletion of interval i400 partially disrupts spore killing but does not eliminate it. Future work should retest the i400 strains in spore killing assays, to determine if the partial spore killing phenotype can be detected in all crosses when larger numbers of rosettes are examined. The findings presented here help narrow down the search for the unknown poison gene involved in spore killing, which is a critical step towards understanding processes that allow for the evolution of Sk-3-type selfish genetic elements.  more » « less
Award ID(s):
2005295
PAR ID:
10649513
Author(s) / Creator(s):
Publisher / Repository:
ISU ReD: Research and eData
Date Published:
Subject(s) / Keyword(s):
Senior Thesis
Format(s):
Medium: X
Institution:
Illinois State University
Sponsoring Org:
National Science Foundation
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  1. (, ISU ReD: Research and eData)
    Meiotic drive elements, sometimes called selfish genes, are genetic elements that are passed on to their offspring more favorably than other genes. Meiotic drive elements have been observed in many organisms, including fungi of the Neurospora genus. Three different meiotic drive elements, called Spore killers, have been identified in Neurospora fungi. One of these Spore killers is called Spore killer 3 (Sk-3), and the molecular mechanism by which Sk-3 acts as a meiotic drive element is poorly understood. Previous work has identified a genetic locus within Sk-3 that may control spore killing. In this thesis, I investigate an interval of DNA within this locus called v375. Through gene deletion and spore killing assays, I show that deletion of v375 disrupts Sk-3-based spore killing in N. crassa. Possible explanations for why interval v375 is required for spore killing by Sk-3 are discussed. 
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  2. (, ISU ReD: Research and eData)
    Neurospora crassa is a fungus that serves as a model organism for genetic research. N. crassa Spore killer-3 (Sk-3) is a genetic element transmitted to offspring through spore killing. Sk-3 is located on Chromosome III and it is thought to require two genes for spore killing. These two genes are the poison gene, for killing, and the antidote gene, for resistance to killing. While the Sk-3 resistance gene has been identified (rsk), the Sk-3 killer gene has not. The primary goal of this study is to help identify the killer gene by investigating the role of a DNA interval called v377 in spore killing. To determine if this interval is required for spore killing, a DNA deletion vector (Vector v377) was constructed and used to replace the v377 interval with a hygromycin resistance gene in strain RDGR170.3. My results demonstrate that v377 is required for spore killing. The possibility that v377 is within a gene required for spore killing, or a regulatory element that controls spore killing, is discussed. 
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  3. (, ISU ReD: Research and eData)
    Meiotic drive describes a process in which selfish alleles are recovered in more than half of a progeny generation. It is a type of gene drive and it has been discovered in strains of Neurospora, a filamentous fungus, through its spore killing mechanism. One of the most studied meiotic drive elements within N. crassa is Spore killer-3 (Sk-3). Previous studies have indicated that there is a genomic region within Sk-3 that encodes resistance to spore killing and another that encodes an element that is required for spore killing. Sk-3’s resistance gene, rsk, has been identified. However, the exact region that mediates Sk-3’s spore killing mechanism is currently unknown. In a previous study, it was found that a mutation called rfk-2UV disrupts spore killing by Sk-3. To better understand the region of Chromosome III in which rfk-2UV is located (its exact location is unknown), I constructed a deletion vector to replace a DNA interval (v374) with a hygromycin resistance gene marker (hph). Transformants were crossed to produce offspring, and offspring were tested to determine if they possess the ability to kill ascospores. These findings will contribute to future efforts to determine the molecular nature of rfk-2UV and why this mutation disrupts the ability of Sk-3 to kill spores. 
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  4. (, ISU ReD: Research and eData)
    Some isolates of the fungus Neurospora crassa possess a chromosomal factor that causes spore killing, leading to death of ascospores. It has been shown that these chromosomal factors are genetic elements called spore killers. For example, if a cross is performed between a parent with an Sk-S (sensitive) allele and a parent with an Sk-K (killer) allele, the cross will produce half viable offspring and half inviable offspring, where the inviable half has been killed by spore killing. This phenomenon can be explained by meiotic drive, wherein a selfish gene disrupts the randomness of sexual transmission, favoring its own success. In this study, I focus on a Neurospora Spore killer known as Sk-3. Sk-3 is thought to possess both a killer element and a resistance element. The resistance element is rsk, a gene that keeps ascospores alive and viable when in the presence of the killer element. However, the mechanism by which the killer element kills ascospores is unknown. A major obstacle to studying the killing mechanism is that the identity of the Sk-3 killer element itself has remained elusive. My goal is to help identify the Sk-3 killer element. Preliminary results by others have narrowed the search to the left arm of Chromosome III. These results have also shown that deletion of a 1.3 kb DNA interval, called v350, causes loss of spore killing. This suggests that a regulatory element, or a hidden gene, may overlap with the v350 interval. To help determine why v350 deletion correlates with loss of spore killing, I investigated a related DNA interval, called v384. My results suggest that v384, like v350, is required for spore killing. 
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  5. (, ISU ReD)
    Neurospora fungi are found around the world. The species N. crassa is a popular model for use in genetics research. N. crassa produces sexual spores, called ascospores, during mating between strains of opposite mating types. N. crassa also produces spore sacs called asci, and each ascus typically contains eight viable ascospores. However, some Neurospora fungi carry selfish genetic elements called Spore killers, and when a strain carrying a Spore killer mates with a spore killing-susceptible strain, asci contain four black viable ascospores and four white inviable ascospores. In this project, I investigated a Spore killer called Sk-3. To act as a selfish genetic element, Sk-3 is thought to require at least two genes, a poison gene and an antidote gene. The Sk-3 antidote gene (rsk) has been identified, but the poison gene has not. The purpose of this study is to help identify the location of the poison gene. To do this, I deleted two DNA intervals (i383 and i394) from a location of the Neurospora genome that may harbor the poison gene. My results indicate that deletion of i383 eliminates spore killing while deletion of i394 has no effect on spore killing. The possibility that i394 overlaps with the poison gene is discussed. 
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