TY - JOUR
T1 - The Patterning and Proportion of Charged Residues in the Arginine-Rich Mixed-Charge Domain Determine the Membrane-Less Organelle Targeted by the Protein
AU - Miyagi, Tamami
AU - Yamazaki, Rio
AU - Ueda, Koji
AU - Narumi, Satoshi
AU - Hayamizu, Yuhei
AU - Uji-i, Hiroshi
AU - Kuroda, Masahiko
AU - Kanekura, Kohsuke
N1 - Funding Information:
This work was financially supported by the JSPS KAKENHI (20H03593 to K.K., 21H02706 to M.K., 21H04634 to H.U., 19H03627 to S.N., 20H02564 to Y.H.).
Publisher Copyright:
© 2022 by the authors.
PY - 2022/7
Y1 - 2022/7
N2 - Membrane-less organelles (MLOs) are formed by biomolecular liquid–liquid phase separation (LLPS). Proteins with charged low-complexity domains (LCDs) are prone to phase separation and localize to MLOs, but the mechanism underlying the distributions of such proteins to specific MLOs remains poorly understood. Recently, proteins with Arg-enriched mixed-charge domains (R-MCDs), primarily composed of R and Asp (D), were found to accumulate in nuclear speckles via LLPS. However, the process by which R-MCDs selectively incorporate into nuclear speckles is unknown. Here, we demonstrate that the patterning of charged amino acids and net charge determines the targeting of specific MLOs, including nuclear speckles and the nucleolus, by proteins. The redistribution of R and D residues from an alternately sequenced pattern to uneven blocky sequences caused a shift in R-MCD distribution from nuclear speckles to the nucleolus. In addition, the incorporation of basic residues in the R-MCDs promoted their localization to the MLOs and their apparent accumulation in the nucleolus. The R-MCD peptide with alternating amino acids did not undergo LLPS, whereas the blocky R-MCD peptide underwent LLPS with affinity to RNA, acidic poly-Glu, and the acidic nucleolar protein nucleophosmin, suggesting that the clustering of R residues helps avoid their neutralization by D residues and eventually induces R-MCD migration to the nucleolus. Therefore, the distribution of proteins to nuclear speckles requires the proximal positioning of D and R for the mutual neutralization of their charges.
AB - Membrane-less organelles (MLOs) are formed by biomolecular liquid–liquid phase separation (LLPS). Proteins with charged low-complexity domains (LCDs) are prone to phase separation and localize to MLOs, but the mechanism underlying the distributions of such proteins to specific MLOs remains poorly understood. Recently, proteins with Arg-enriched mixed-charge domains (R-MCDs), primarily composed of R and Asp (D), were found to accumulate in nuclear speckles via LLPS. However, the process by which R-MCDs selectively incorporate into nuclear speckles is unknown. Here, we demonstrate that the patterning of charged amino acids and net charge determines the targeting of specific MLOs, including nuclear speckles and the nucleolus, by proteins. The redistribution of R and D residues from an alternately sequenced pattern to uneven blocky sequences caused a shift in R-MCD distribution from nuclear speckles to the nucleolus. In addition, the incorporation of basic residues in the R-MCDs promoted their localization to the MLOs and their apparent accumulation in the nucleolus. The R-MCD peptide with alternating amino acids did not undergo LLPS, whereas the blocky R-MCD peptide underwent LLPS with affinity to RNA, acidic poly-Glu, and the acidic nucleolar protein nucleophosmin, suggesting that the clustering of R residues helps avoid their neutralization by D residues and eventually induces R-MCD migration to the nucleolus. Therefore, the distribution of proteins to nuclear speckles requires the proximal positioning of D and R for the mutual neutralization of their charges.
KW - liquid–liquid phase separation
KW - membrane-less organelle
KW - nuclear speckle
KW - nucleolus
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U2 - 10.3390/ijms23147658
DO - 10.3390/ijms23147658
M3 - Article
C2 - 35887012
AN - SCOPUS:85135127677
SN - 1661-6596
VL - 23
JO - International Journal of Molecular Sciences
JF - International Journal of Molecular Sciences
IS - 14
M1 - 7658
ER -