35S rRNA transcripts include a 5'-external transcribed spacer followed by rRNAs of the small and large ribosomal subunits. Their processing yields massive precursors that include dozens of assembly factor proteins. In Saccharomyces cerevisiae, nucleolar assembly factors form 2 coaxial layers/volumes around ribosomal DNA. Most of these factors are cyclically recruited from a latent state to an operative state, and are extensively conserved. The layers match, at least approximately, known subcompartments found in higher eukaryotic cells. ∼80% of assembly factors are essential. The number of copies of these assembly factors is comparable to the number of nascent transcripts. Moreover, they exhibit "isoelectric balance," with RNA-binding candidate "nucleator" assembly factors being notably basic. The physical properties of pre-small subunit and pre-large subunit assembly factors are similar, as are their 19 motif signatures detected by hierarchical clustering, unlike motif signatures of the 5'-external transcribed spacer rRNP. Additionally, many assembly factors lack shared motifs. Taken together with the progression of rRNP composition during subunit maturation, and the realization that the ribosomal DNA cable is initially bathed in a subunit-nonspecific assembly factor reservoir/microenvironment, we propose a "3-step subdomain assembly model": Step (1): predominantly basic assembly factors sequentially nucleate sites along nascent rRNA; Step (2): the resulting rRNPs recruit numerous less basic assembly factors along with notably basic ribosomal proteins; Step (3): rRNPs in nearby subdomains consolidate. Cleavages of rRNA then promote release of rRNPs to the nucleoplasm, likely facilitated by the persistence of assembly factors that were already associated with nucleolar precursors.

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Journal Article | Research Support, N.I.H., Extramural | Research Support, Non-U.S. Gov't

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Lin S, Rajan S, Lemberg S, Altawil M, Anderson K, Bryant R, Cappeta S, Chin B, Hamdan I, Hamer A, ... Show all

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