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Molecular partnership of the intrinsically disorder C-terminal domain of the measles virus nucleoprotein (NTAIL)

Heads Sonia LONGHI

  Assessing the functional impact of the Measles virus NTAIL-PXD interaction in vivo

Molecular partnership of the intrinsically disorder C-terminal domain of the measles virus nucleoprotein (NTAIL)

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Figure 5.
(A) (Top) Structural organization of the N protein showing that it consists of an ordered amino terminal NCORE domain and an intrinsically disordered C-terminal NTAIL domain. The three NTAIL regions (referred to as Box-1, 2 and 3) conserved among Morbillivirus members are shown, where PXD binds Box-2. (Bottom) The sequence of Box-2 is shown along with the amino acid substitutions that were tested in the analysis of PXD/NTAIL structural and functional interactions. The predicted α-MoRE and the PXD interacting sequence are shown. The region adopting an α–helical conformation in the NTAIL-PXD complex spans amino acids 486-502. (B) Lateral and (C) end-on views of the structure of PXD in complex with residues 486-504 of NTAIL. The side chains of the substituted amino acids (displayed in sticks) are shown to illustrate their relative orientation with respect to PXD. Modified from (Shu et al., J Biol Chem 2012).

People involved: J. Habchi (PhD student)

The MeV P protein tethers the polymerase to the nucleocapsid template for transcription and genome replication. Binding of P to nucleocapsid is mediated by the X domain of P and a conserved sequence (Box2, aa 489-506) within the C-terminal domain of the nucleoprotein (NTAIL). PXD binding induces NTAIL α-helical folding, which in turn has been proposed to stabilize the polymerase-nucleocapsid complex, with cycles of binding and release required for transcription and genome replication. In a recent study carried out in collaboration with the groups of M. Oglesbee (Ohio State University) and of D. Gerlier (Lab. Virologie Humaine, Lyon), we assessed the relationships between PXD-induced NTAIL folding, XD-NTAIL binding affinity, and polymerase activity. Amino acid substitutions that abolished PXD-induced NTAILTAIL (Figure 5). Surprisingly, polymerase activity in minireplicons was maintained despite a 35-fold decrease in PXD/ NTAIL binding affinity or reduction/loss of PXD-induced NTAIL folding. Recombinant infectious virus was recovered for all mutants, and transcriptase elongation rates remained within a 1.7-fold range of parent virus. Box2 substitutions did however impose a significant cost to infectivity, reflected in an increase in the amount of input genome required to match the infectivity of parent virus. Diminished infectivity could not be attributed to changes in virion protein composition or production of defective interfering particles, where changes from parent virus were within a 3-fold range. Results indicate that MeV polymerase activity, but not infectivity, tolerates amino acid changes in the PXD-binding region of the nucleoprotein. Selectional pressure for conservation of Box2 sequence may thus reflect a role in assuring the fidelity of polymerase functions or the assembly of viral particles required for optimal infectivity.

Collectively, these studies provided a new perspective for the study of viral polymerase activity and its modulation by viral and host factors. In particular, the currently accepted model whereby the NTAIL-PXD interaction has to be relatively weak in order to allow the polymerase to cartwheel on the nucleocapsid template needs to be revisited in light of the present results.

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