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At the Anterior: acts as a TF to aid in REPRESSION of caudal mRNA TransLATION
Process of inhibiting transLATION of Caudal mRNA
1. 5' caudal (cad) cap binding by d4EHP (protein) + 3' cad binding by Bicoid bound to BBR: both act together to block translation
Bicoid Main points
- 1. d4EHP and del4FE have similar structures, except for a few tyrosines
- 2. d4EHP mutants made, showed that Bcd binds to d4EHP, at dorsal end, as when mutant at Trp85, the binding was gone. (Fig 3)
- 3. d4EHP interacts with BOTH Bcd and cad mRNA 5' to inhibit cad expression, as when mutants at 85 and 114, the CAD expression EXTENDS TO ANTERIOR, no gradient as in WT (Fig 4)
- 4. the d4EHP interaction with Bcd requires a sequence motif that is DIFFERENT than the one in del4FE or 4E-BP (Fig 5)
- 5. when implanting these mutations of Bcd found from Fig 5 (66 and 73), the Cad repression was NOT seen, and the embryos mostly failed to hatch, and the ones that hatched failed to develop into adults (Fig 6)
- 6. Bcd and d4EHP interaction is needed to INHIBIT TransLATION, as when they made a rLuc construct with cad mRNA (BBRsense), then translated with presence of d4EHP, repression seen in rLuc vs when the BBRantisense was used (Fig 7)
- 7. Mechanism for inhibition of Translation: Bcd interacts with d4EHP so that both the 3' UTR (via BBR) and the 5' cap are bound, respectively, to inhibit translation of CaudalmRNA
- elF4E binding proteins (E4-BP) bring elF4E to bind at the 5' cap of caudal mRNA. Competition occurs btw elF4G for binding of elF4E.
- Bicoid aids in blocking translation by interacting with elF4E. Bicoid competes with elF4G to bind to d4EHP.
d4EHP acts to compete with the E4-BP protein complex for capping of the 5' caudalmRNA, and Bicoid binds to the d4EHP at specific residues to aid in inhibition of translation by binding its BBR to the 3' UTR of the cad mRNA
Bicoid experiments done:
- Fig 1.
- d4EHP and delF4E have similar binding cap structures, except for 2 tyrosines (43 and 56)
- d4EHP and delF4E both bind to GTP Sephorase
- Fig 2.
- a. Mutant d4EHP cp53 mutant shown, with one intron removed
- b. mutant still has some d4EHP expression
- c. seen at reduced levels vs WT
- d. 12 amino acids missing vs WT
- e. IMMUNOFLUORESCENCE shows D4EHP mutant shows ANTERIOR Cad expression, although similar zygotic hb activation and cad localization vs WT.
- Fig 3.
- Summary: d4EHP interacts with Bcd in vitro and in vivo
- Fig 4.
- d4EPH mutations at 58 and 85 NOT able to INHIBIT translation of ANTERIOR cad mRNA