Insect Elytra Formation
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Questions and Answers

What are the two major cuticular proteins identified in the study?

  • TcCPR27 and TcCPR30
  • TcCPR22 and TcCPR18
  • TcCPR15 and TcCPR20
  • TcCPR27 and TcCPR18 (correct)

    • What phenotype did TcCPR27-deficient insects exhibit?

  • Smooth elytra with normal folding
  • Shorter, wrinkled, and warped elytra (correct)
  • Longer, robust elytra
  • No significant changes in elytra structure

    • At which life stage did transcripts for TcCPR27 and TcCPR18 increase dramatically?

  • Pupal stage
  • Pharate adult stage (correct)
  • Larval stage
  • Embryonic stage

    • What was the outcome for TcCPR27-deficient insects shortly after eclosion?

    <p>They died prematurely, likely due to dehydration.</p> Signup and view all the answers

    What does the elytra of a beetle serve as?

    <p>A highly sclerotized cover for the hindwings</p> Signup and view all the answers

    Which family do TcCPR27 and TcCPR18 belong to?

    <p>Rebers and Riddiford family</p> Signup and view all the answers

    What method was employed to examine the presence of TcCPR27 in the elytral cuticle?

    <p>Immunolocalization studies</p> Signup and view all the answers

    What was the effect of injecting double-stranded RNAs targeting TcCPR27 and TcCPR18 on larval development?

    <p>There was no effect on larval to pupal transformation.</p> Signup and view all the answers

    What is the primary function of TcCPR27 and TcCPR18 in the Tribolium species?

    <p>To provide rigidity to the cuticle</p> Signup and view all the answers

    During which stage do transcript levels of TcCPR27 and TcCPR18 dramatically increase?

    <p>At the pharate adult stage</p> Signup and view all the answers

    Which statement is true regarding the expression of TcCPR27 and TcCPR18 in larvae?

    <p>Few or no transcripts are detected during this stage.</p> Signup and view all the answers

    What do the proteins extracted from the pronotum and elytra after eclosion indicate?

    <p>Presence of rigidity proteins post-eclosion.</p> Signup and view all the answers

    What was observed about TcCRP27 localization in pharate adults?

    <p>It co-localized with chitin in the dorsal elytral cuticle.</p> Signup and view all the answers

    What happens to the cuticle in the pronotum and ventral abdomen as the adult matures?

    <p>It becomes more rigid and darker.</p> Signup and view all the answers

    How do the transcript levels of TcCPR27 and TcCPR18 compare in the elytron and hindwing?

    <p>They are significantly higher in the elytron.</p> Signup and view all the answers

    What was the effect of injecting dsRNA for TcLac2 into day 0 pupae?

    <p>It suppressed cuticle tanning.</p> Signup and view all the answers

    What does E9 measure regarding the elytra?

    <p>The stiffness of the elytra</p> Signup and view all the answers

    What effect did the injection of dsTcCPR27 have on the elytra?

    <p>Decreased rigidity</p> Signup and view all the answers

    Which protein was indicated to have a more severe phenotype upon knockdown?

    <p>TcCPR27</p> Signup and view all the answers

    How does maintaining a constant concentration of quinone cross-linking molecules affect protein molecules?

    <p>It leads to a greater average number of cross-links per protein molecule</p> Signup and view all the answers

    What was the purpose of injecting dsVer in the experiments?

    <p>To serve as a negative control</p> Signup and view all the answers

    What is a possible reason for the different severity in phenotypes between TcCPR27 and TcCPR18?

    <p>Difference in their structural properties</p> Signup and view all the answers

    Which species were expressed in protein extracts similar to TcCPR27 and TcCPR18?

    <p>Three other Tribolium species</p> Signup and view all the answers

    What is the role of quinone cross-linking molecules in the context of cuticular proteins?

    <p>To stabilize the cuticle structure</p> Signup and view all the answers

    What was the primary observation regarding the removal of the hindwings in T. castaneum?

    <p>It had no effect on adult survival.</p> Signup and view all the answers

    What is the theoretical molecular mass of TcCPR27?

    <p>11.4 kDa</p> Signup and view all the answers

    Which amino acid does TcCPR18 replace in its RR2 motif?

    <p>Glycine</p> Signup and view all the answers

    What was the consequence of removing the elytra from adult T. castaneum?

    <p>It caused high mortality due to dehydration.</p> Signup and view all the answers

    Which of the following proteins was predominant in the elytra of various Tribolium species?

    <p>TmACP22</p> Signup and view all the answers

    What indicates that TcCPR27 and TcCPR18 had not undergone substantial cross-linking?

    <p>The high yields in extracts</p> Signup and view all the answers

    What was identified as the possible amino-terminal residue of the mature TcCPR27 protein?

    <p>Pyroglutamic acid</p> Signup and view all the answers

    What is the primary difference in cuticle properties between TcCPR27 and TcCPR18 with respect to their localization?

    <p>TcCPR27 and TcCPR18 are major proteins in highly sclerotized cuticles</p> Signup and view all the answers

    How were the major proteins in the elytra analyzed?

    <p>Through 4–12% Bis-Tris gel electrophoresis.</p> Signup and view all the answers

    In the context of their sequence identity, what is true about TcCPR18?

    <p>It is an ortholog of a protein in Tenebrio molitor</p> Signup and view all the answers

    What is the observed mass of the native 10 kDa band determined by MALDI-linear-TOF MS for TcCPR27?

    <p>11,467 Da</p> Signup and view all the answers

    What characteristic is associated with TcCPR18 in comparison to TcCPR27?

    <p>Higher histidine content</p> Signup and view all the answers

    What effect does the eclosion timing have on the extraction yields of TcCPR27 and TcCPR18?

    <p>Higher yields in newly emerged adults</p> Signup and view all the answers

    What type of protein are TmACP20 and TmACP22 identified as?

    <p>Cuticular proteins</p> Signup and view all the answers

    What were the apparent masses of the two to three abundant proteins obtained from each Tribolium species?

    <p>10 and 20 kDa</p> Signup and view all the answers

    What happens to the dorsal abdominal cuticle in adults as it relates to TcCPR27 and TcCPR18?

    <p>It remains flexible and membranous</p> Signup and view all the answers

    What process was used to remove protein from the elytra before staining?

    <p>Incubation with 10 N NaOH at 95°C</p> Signup and view all the answers

    What type of staining was performed on the elytra?

    <p>Fluorescein-conjugated chitin-binding domain staining</p> Signup and view all the answers

    What was the purpose of injecting dsRNA into the pupae?

    <p>To investigate the role of specific genes in development</p> Signup and view all the answers

    Who performed the gene cloning, protein purification, and immunohistochemistry?

    <p>YA</p> Signup and view all the answers

    What type of analysis was performed by JL?

    <p>Mechanical analysis</p> Signup and view all the answers

    What was injected at the late larval instar stage?

    <p>dsRNA for specific genes</p> Signup and view all the answers

    What is the significance of the elytra appearance not differing until after adult eclosion?

    <p>It indicates the process of cuticle sclerotization is complete.</p> Signup and view all the answers

    What can be inferred about the techniques used in this study?

    <p>They integrate molecular biology and biochemical methods.</p> Signup and view all the answers

    Study Notes

    Beetle Elytra Formation

    • Beetle forewings (elytra) are modified for protection of hindwings and abdomen
    • They are rigid and sclerotized (hardened)
    • Two major cuticular proteins (TcCPR27 and TcCPR18) are crucial for elytra formation
    • These proteins are abundant in pronotum and ventral abdominal cuticles
    • They belong to the Rebers and Riddiford family of cuticular proteins
    • Contain RR2 motifs, potentially involved in chitin binding
    • Expression dramatically increases at pharate adult stage and then decreases
    • Injecting dsRNA for these proteins into larvae results in shorter, wrinkled, warped, fenestrated elytra, less rigid than controls
    • TcCPR27 deficiency leads to premature death (approx 1 week after eclosion) due to dehydration, TcCPR18 deficiency shows similar but less severe phenotype
    • TcCPR27 is found within the elytral cuticle
    • Both proteins are required for correct elytra morphogenesis

    Cuticle Composition and Function

    • Insect cuticle is primarily chitin and structural proteins.
    • The epicuticle provides waterproofing.
    • The procuticle consists of protein-rich layers and chitin-rich procuticle layers
    • Cuticular proteins (CPs) and chitin are the primary structural components of the procuticle.
    • Post-translational modifications like sclerotization and pigmentation harden the cuticle for protection
    • Insect CPs form various families with specific motifs
    • CPR family is the largest, containing the Rebers & Riddiford (R&R) motif likely involved in chitin binding

    Protein Expression and Localization

    • TcCPR27 and TcCPR18 are abundant in rigid cuticles (elytra, pronotum, ventral abdomen)
    • Expression is low or absent in flexible cuticles (dorsal abdomen, hindwings)
    • Expression dramatically increases at the pharate adult stage and rapidly decreases
    • Proteins are not detected during egg, larval, and early pupal stages
    • TcCPR27 protein is localized in dorsal elytra and ventral abdomen, correlated with chitin
    • Immunostaining confirms localization

    RNAi and Phenotype Analysis

    • RNA interference (RNAi) was used to knock down TcCPR27 and TcCPR18 expression
    • RNAi knockdown of these proteins caused elytral malformation (short, wrinkled, warped, fenestrated) and hindwing folding problems
    • TcCPR27 deficiency resulted in more significant morphological defects compared to TcCPR18

    Mechanical Analysis

    • Dynamic mechanical analysis (DMA) of elytra showed that TcCPR27-deficient elytra were significantly less rigid than controls
    • Downregulation of TcCPR27 resulted in lower elastic modulus (E') and lower tan δ, suggesting more cross-linking of the elytral cuticle.

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    Related Documents

    Beetle Forewing Formation Requires Two Major Proteins (2012) PDF

    Description

    This quiz explores the formation and function of beetle elytra, highlighting the roles of key proteins TcCPR27 and TcCPR18. Learn about the structural adaptations of beetle forewings and their significance in insect morphology and protection. Test your knowledge on how these proteins influence elytral formation and insect survival.

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