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Poster De Conférence Année : 2021

Structural analysis of the composite mitochondrial ribosome assembly factor YBEY-p32

Résumé

YBEY is a small ultraconserved protein with endoribonuclease activity found in most bacteria, mitochondria and chloroplasts. It plays a key role in the ribosome biogenesis and is, therefore, critical for cell physiology. We have previously shown that human YBEY does not participate in rRNA processing and instead helps to incorporate the ribosomal protein uS11m into the nascent small subunit of the mitochondrial ribosome, which is required for the organellar translation and cellular respiration. Unlike its bacterial orthologues, human YBEY exists in a stable complex with a deeply conserved mitochondrial trimeric protein p32 (a.k.a. C1QBP). Similarly to YBEY, p32/C1QBP is essential for mitochondrial translation, albeit its molecular mechanism has not been established. C1QBP mutations cause mitochondrial diseases, whereas its overexpression is a hallmark of aggressive cancers. We found that the YBEY-p32 complex stoichiometrically recruits uS11m, which likely plays an important role in its delivery to the ribosomes. However, the molecular details of this unusual composite assembly factor and the exact contribution of p32 to the YBEY-uS11m pathway remain unclear. Here we show that, unlike bacterial YbeY proteins (usually negatively charged), the human YBEY homologue is neutral (pI = 7.04) and, as a consequence, poorly soluble in isolation. However, its association with the highly acidic p32 (pI ≈ 4) compensates for this deficiency and communicates both solubility and an overall negative charge to the particle. This property might be important for its interaction with the positively charged uS11m. To establish which YBEY residues are involved in p32 binding, we created a collection of YBEY variants with point mutations in conserved residues and systematically evaluated their ability to form a complex with p32 upon coexpression in E. coli. We found that specific amino acids at and near the presumed active centre of YBEY are critical for its association with p32. Due to this specific binding topology, p32 sterically constrains the active centre of YBEY and inhibits its intrinsic RNase activity without compromising its ability to interact with uS11m. We propose that the human YBEY-p32 complex functions primarily via uS11m binding and is unlikely to be an RNase in vivo.
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Dates et versions

hal-03253326 , version 1 (08-06-2021)

Identifiants

  • HAL Id : hal-03253326 , version 1

Citer

Cédric Schelcher, Christelle Gruffaz, Alexandre Smirnov. Structural analysis of the composite mitochondrial ribosome assembly factor YBEY-p32. The 26th Annual Meeting of the RNA Society - RNA 2021, May 2021, On-line, United States. . ⟨hal-03253326⟩
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