Date Thesis Awarded
5-2024
Access Type
Honors Thesis -- Access Restricted On-Campus Only
Degree Name
Bachelors of Science (BS)
Department
Biology
Advisor
Diane C. Shakes
Committee Members
Oliver Kerscher
Giulia Pacini
Abstract
Unlike human flagellated sperm, C. elegans sperm use the movement of an extended pseudopod to reach and persist in the site of fertilization. This research uses a knockout mutant (spe-54) to examine the role of the protein SPE-54 in C. elegans sperm.
To determine the impact of spe-54 on C. elegans male and hermaphrodite fertility, the success of male fertility between the mutant spe-54 and control strains was measured. The quantification of viable progeny and unfertilized oocytes showed decreased spe-54 fertility compared to him-5 (a wild-type nematode) in both male fertility and hermaphrodite fertility experiments.
To determine the cause of reduced fertility in the spe-54 mutant, the distribution of fluorescent sperm was observed in the reproductive tract of fog-2 (mutant nematode) females following insemination by either spe-54 or him-5 males. DIC/fluorescent images show that spe-54 sperm fail to migrate to the site of fertilization, causing unfertilized oocytes to be lain.
To investigate the cause of reduced sperm movement, sperm were labeled with anti-MSP to determine the localization of MSP (Major Sperm Protein), a structural protein used for cell motility localized in the sperm pseudopod. Immunofluorescence images show a shortened, broadened MSP localization (crescent shape) compared to the extended rectangle/triangle shape of the control MSP localization.
To determine the impact of a short pseudopod in sperm movement, the treadmilling (cyclic pseudopodal motion) rates of spe-54 sperm and him-5 controls were quantified. In general, the shortened pseudopod of spe-54 led to decreased treadmilling rates compared to control sperm. Thus, the shortened pseudopod contributes to lack of motility and subsequent failure to reach the site of fertilization in the reproductive tract.
Further immunofluorescent analyses to determine how SPE-54 regulates sperm function produced different localization patterns of both SPE-6 and SPE-9 proteins between spe-54 and control sperm.
Extension of a pseudopod and the fusion of Membranous Organelles (MOs) are two key features of C. elegans sperm activation. To determine if spe-54 impacts the process of sperm activation, a mutant that lacks MO fusion and an extended pseudopod (fer-1) was compared to spe-54 Using a membrane dye (FM1-43), fer-1 strains showed no MO fusion while both him-5 and spe-54 showed frequent presence of fused MOs. Thus, spe-54, while lacking pseudopodal extension, still retains MO fusion in the sperm activation process.
In all, the lack of spe-54 is shown to lead to altered pseudopod shape and decreased treadmilling rates in C. elegans sperm. These findings provide the explanation of overall decreased fertility in the spe-54 mutant strains.
Recommended Citation
Vanella, Corinne, "spe-54 required for directed cell motility in Caenorhabditis elegans sperm" (2024). Undergraduate Honors Theses. William & Mary. Paper 2221.
https://scholarworks.wm.edu/honorstheses/2221
