Ferlins are membrane proteins with multiple C2 domains and proposed functions in Ca2+ mediated me... more Ferlins are membrane proteins with multiple C2 domains and proposed functions in Ca2+ mediated membrane-membrane interactions in animals. Caenorhabditis elegans has two ferlin genes, one of which is required for sperm function. Mammals have several ferlin genes and mutations in the human dysferlin (DYSF) and otoferlin (OTOF) genes result in muscular dystrophy and hearing loss, respectively. Drosophila melanogaster has a single ferlin gene called misfire (mfr). A previous study showed that a mfr mutation caused male sterility because of defects in fertilization. Here we analyze the expression and structure of the mfr gene and the consequences of multiple mutations to better understand the developmental function of ferlins. We show that mfr is expressed in the testis and ovaries of adult flies, has tissue-specific promoters, and expresses alternatively spliced transcripts that are predicted to encode distinct protein isoforms. Studies of 11 male sterile mutations indicate that a predicted Mfr testis isoform with five C2 domains and a transmembrane (TM) domain is required for sperm plasma membrane breakdown (PMBD) and completion of sperm activation during fertilization. We demonstrate that Mfr is not required for localization of Sneaky, another membrane protein necessary for PMBD. The mfr mutations vary in their effects in females, with a subset disrupting egg patterning and causing a maternal effect delay in early embryonic development. Locations of these mutations indicate that a short Mfr protein isoform carries out ferlin activities during oogenesis. The mfr gene exhibits complex transcriptional and post-transcriptional regulation and functions in three developmental processes: sperm activation, egg patterning, and early embryogenesis. These functions are in part due to the production of protein isoforms that vary in the number of C2 domains. These findings help establish D. melanogaster as model system for understanding ferlin function and dysfunction in animals, including humans.
Undergraduate students in genetics and developmental biology courses often struggle with the conc... more Undergraduate students in genetics and developmental biology courses often struggle with the concept of epistasis because they are unaware that the logic of gene interactions differs between enzymatic pathways and signaling pathways. If students try to develop and memorize a single simple rule for predicting epistatic relationships without taking into account the nature of the pathway under consideration, they can become confused by cases where the rule does not apply. To remedy this problem, we developed a short pre-/post-test, an in-class activity for small groups, and a series of clicker questions about recessive epistasis in the context of a signaling pathway that intersects with an enzymatic pathway. We also developed a series of homework problems that provide deliberate practice in applying concepts in epistasis to different pathways and experimental situations. Students show significant improvement from pretest to posttest, and perform well on homework and exam questions following this activity. Here we describe these materials, as well as the formative and summative assessment results from one group of students to show how the activities impact student learning.
Ferlins are membrane proteins with multiple C2 domains and proposed functions in Ca2+ mediated me... more Ferlins are membrane proteins with multiple C2 domains and proposed functions in Ca2+ mediated membrane-membrane interactions in animals. Caenorhabditis elegans has two ferlin genes, one of which is required for sperm function. Mammals have several ferlin genes and mutations in the human dysferlin (DYSF) and otoferlin (OTOF) genes result in muscular dystrophy and hearing loss, respectively. Drosophila melanogaster has a single ferlin gene called misfire (mfr). A previous study showed that a mfr mutation caused male sterility because of defects in fertilization. Here we analyze the expression and structure of the mfr gene and the consequences of multiple mutations to better understand the developmental function of ferlins. We show that mfr is expressed in the testis and ovaries of adult flies, has tissue-specific promoters, and expresses alternatively spliced transcripts that are predicted to encode distinct protein isoforms. Studies of 11 male sterile mutations indicate that a predicted Mfr testis isoform with five C2 domains and a transmembrane (TM) domain is required for sperm plasma membrane breakdown (PMBD) and completion of sperm activation during fertilization. We demonstrate that Mfr is not required for localization of Sneaky, another membrane protein necessary for PMBD. The mfr mutations vary in their effects in females, with a subset disrupting egg patterning and causing a maternal effect delay in early embryonic development. Locations of these mutations indicate that a short Mfr protein isoform carries out ferlin activities during oogenesis. The mfr gene exhibits complex transcriptional and post-transcriptional regulation and functions in three developmental processes: sperm activation, egg patterning, and early embryogenesis. These functions are in part due to the production of protein isoforms that vary in the number of C2 domains. These findings help establish D. melanogaster as model system for understanding ferlin function and dysfunction in animals, including humans.
Undergraduate students in genetics and developmental biology courses often struggle with the conc... more Undergraduate students in genetics and developmental biology courses often struggle with the concept of epistasis because they are unaware that the logic of gene interactions differs between enzymatic pathways and signaling pathways. If students try to develop and memorize a single simple rule for predicting epistatic relationships without taking into account the nature of the pathway under consideration, they can become confused by cases where the rule does not apply. To remedy this problem, we developed a short pre-/post-test, an in-class activity for small groups, and a series of clicker questions about recessive epistasis in the context of a signaling pathway that intersects with an enzymatic pathway. We also developed a series of homework problems that provide deliberate practice in applying concepts in epistasis to different pathways and experimental situations. Students show significant improvement from pretest to posttest, and perform well on homework and exam questions following this activity. Here we describe these materials, as well as the formative and summative assessment results from one group of students to show how the activities impact student learning.
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