BIOL 411 - 2023 Autumn - Exam 1 - Answer Key

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BIOL 411 - 2023 Autumn Exam 1 Point total: 100 points Instructions (please read carefully): ● Begin by downloading this document (File > Download > Microsoft Word (.docx)) ● Using your own words, write precisely and concisely. Do not exceed the sentence limits. ● Reminder: you may refer to your one page of notes but no other sources during the exam period. ● Unless specified otherwise, providing multiple answers to a question will result in a loss of points. ● Please make sure to upload your document via Canvas at the end of the exam period. ● Sharing your answers with a peer is strictly prohibited. ● Failure to comply with these guidelines may result in our being unable to accept or grade your exam or initiating disciplinary actions. ● Please format your written answers (bold, colored blue) to make exam grading easier. Integrity statement: I have neither given nor received aid on this exam. _________________________ Your Signature Page 1

BIOL 411 - 2023 Autumn Exam 1 1. Based on the cell lineage shown at right, mark each of the following statements as True or False and provide a one sentence explanation of your reasoning. [4pts each/12pts total] 1a. _ False [2pts] _ The MS cell is specified to become pharyngeal or body wall muscle. ● Reasoning: To test this we would need to isolate the MS cell and place it in a neutral environment (e.g., culture dish) and assay muscle formation, but the lineage was only made based on observations of intact embryos. [2pts] 1b. _ False [2pts] _ The MS cell is determined to become pharyngeal or body wall muscle. ● Reasoning: To test this we would need to isolate the MS cell and place it in a non-neutral environment (e.g., culture dish with other components or different region of the embryo) and assay muscle formation, but the lineage was only made based on observations of intact embryos. [2pts] 1c. _ False [2pts] _ The MS cell is a differentiated pharyngeal or body wall muscle cell. ● Reasoning: False because the lineage indicates MS is a precursor cell that will undergo multiple rounds of cell division before producing differentiated muscle cells (at the tips of the branches). [2pts] Figure legend: An abbreviated embryonic cell lineage of C. elegans that was generated based solely on observations of intact, wild-type embryos. Note that the vertical axis in the diagram represents embryonic time and the branch points represent cell divisions. Individual blastomere names are shown above select branches. 2. You are analyzing embryos produced by adult female flies that are either heterozygous ( nanos/+ ) or homozygous ( nanos/nanos ) for a recessive loss-of-function allele of nanos . For each of these cases, in what proportion of embryos do you expect to see the nanos mutant phenotype (i.e., lack of abdomen)? [4pts each/8pts total] nanos/+ nanos/nanos A. 100% B. 50% C. 25% D. 0% [4pts] E. Cannot determine without paternal genotype A. 100% [4pts] B. 50% C. 25% D. 0% E. Cannot determine without paternal genotype Page 2

BIOL 411 - 2023 Autumn Exam 1 3. As part of Unit 1, you had the opportunity to analyze the landmark paper “ macho-1 encodes a localized mRNA in ascidian eggs that specifies muscle fate during embryogenesis” by Hiroki Nishida and Kaichiro Sawada ( Nature 2001 409:724-729). 3a. This paper makes use of ascidian embryos ( Halocynthia roretzi ) as a model system. You are discussing the paper with a peer who has taken BIOL 355, but not BIOL 411, and is unfamiliar with H. roretzi as a model. You are asked to define three advantages and one disadvantage of H. roretzi as a model for analyzing muscle development compared to mammalian embryos. What do you say? (One complete sentence per bullet point) [2pts each/8pts total] Advantages [6pts] ● Multiple possible answers, e.g.: ○ Defined, invariant cell lineage ○ Externally developing embryos ○ Easy to inject mRNA/oligos ○ Simple body plan with well-described anatomy ○ Cheap, easy to culture in lab, etc. ○ It is a chordate making it closely related to vertebrates Disadvantage [2pts] ● Multiple possible answers, e.g.: ○ Genome may not contain factors relevant for mammalian muscle development ○ Mammalian embryos do not have yellow protoplasm, so mechanisms likely different ○ Great for studying autonomous development, but not the conditional development commonly used in mammalian systems Figure 3 legend: n–r, Embryos at the 110-cell stage fixed and in in situ hybridized with the muscle actin ( HrMA4 ) probe. n, Uninjected control. Actin expression is observed in ten precursor blastomeres of primary muscle cells. o, macho-1 -depleted embryos. p, q, Synthesized macho-1 mRNA was subsequently injected. Actin is ectopically expressed in endoderm ( p ) and epidermis ( q ) blastomeres. r, Subsequent injection of mutant mRNA has no effect. Figure 1 legend: b, Structure of macho-1 protein and constructs for in vitro mRNA synthesis. Macho-1 has five zinc fingers in the central part. The cDNA clone is 2,210 bp long and encodes a 556-amino-acid ORF. The wild-type construct encodes the full length, but the mutant construct lacks the zinc-finger domain. 3b. What do you conclude based on the results presented in Figure 3o ? (Limit: 1 sentence) [4pts] macho-1 is necessary for muscle actin ( HrMA4 ) expression at the 110-cell stage. Page 3

BIOL 411 - 2023 Autumn Exam 1 [1pt] Accurate description of the results. 3c. What do you conclude based on the combined results presented in Figure 3p,q ? (Limit: 1 sentence) [4pts] macho-1 is sufficient to promote muscle actin ( HrMA4 ) expression in endodermal and epidermal blastomeres at the 110-cell stage. [1pt] Accurate description of the results. 3d. Explain to your peer the question the authors were addressing with their mutant mRNA injection. (Limit: 1 sentence) [4pts ] Is injection of macho-1 mRNA lacking the zinc finger domain sufficient to rescue muscle actin ( HrMA4 ) expression in macho-1 -depleted embryos at the 110-cell stage? Alternative answer: the authors were testing if the zinc finger domain is necessary for macho-1 function. [1pt] Accurate description of the results. Argues that it is a rescue experiment or functioning as an injection control. [0pts] The zinc finger domain is not mentioned. Testing if macho-1 mRNA is necessary and sufficient for muscle development was queried in figure 3o,p, and q. 3e. Imagine that injection of the macho-1 mutant mRNA into macho-1 -depleted embryos resulted in embryos that looked like the one presented in Figure 3n . What would you infer about the potential molecular function of Macho-1 protein based on this potential result? (Limit: 1 sentence) [4pts] Macho-1 can promote muscle actin ( HrMA4 ) expression independently of its zinc finger domain. Other possible interpretations: e.g., it doesn’t need to bind to DNA, function as a TF function, etc. to promote muscle actin expression. [2pts] The zinc finger domain is not mentioned but the answer is otherwise correct. 4. During this Unit, you “jigsawed” the seminal paper “The Drosophila Homolog of C. elegans PAR-1 Organizes the Oocyte Cytoskeleton and Directs oskar mRNA Localization to the Posterior Pole” by Joshua M. Shulman, Richard Benton, and Daniel St Johnston ( Cell 2000 101:377–388). 4a. Based on your understanding of the background to this paper, what led the authors to investigate PAR-1 as a candidate regulator of A/P axis determination in Drosophila ? Limit: 1 sentence. [3pts] Previous work in the early C. elegans embryo had identified PAR-1 as an essential regulator of A/P axis determination and the authors wanted to test if this function of PAR-1 was conserved in Drosophila . Page 4

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