Hrr25/CK1δ-directed release of Ltv1 from pre-40S ribosomes is necessary for ribosome assembly and cell growth.
Bottom Line: Conversely, phosphomimetic Ltv1 variants rescued viability after Hrr25 depletion.Finally, Ltv1 knockdown in human breast cancer cells impaired apoptosis induced by CK1δ/ε inhibitors, establishing that the antiproliferative activity of these inhibitors is due, at least in part, to disruption of ribosome assembly.These findings validate the ribosome assembly pathway as a novel target for the development of anticancer therapeutics.
Affiliation: Department of Cancer Biology and Department of Chemistry, The Scripps Research Institute, Jupiter, FL 33458.Show MeSH
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Mentions: The data presented herein, along with previous studies, reveal a two-step mechanism for Rps3 recruitment into the beak structure of pre-40S subunits, where it forms the mRNA entry channel (Fig. 7). Yar1 delivers Rps3 to pre-40S subunits in the nucleus (Koch et al., 2012). Here we show that this is a spontaneous exchange driven by the stronger affinity interaction of Rps3 with pre-40S ribosome–bound Ltv1. In the resulting assembly, intermediate Rps3 is not bound at its mature binding site (Strunk et al., 2011), and can be salt-extracted from pre-40S subunits in a complex with the AFs Enp1 and Ltv1 (Schäfer et al., 2006). Here we show that Hrr25 phosphorylates Ltv1, leading to its release from pre-40S subunits. Enp1 is also released from pre-40S subunits, but the data do not provide strong support for a specific role of Hrr25 in this release. Nevertheless, we do note that Enp1 on pre-40S subunits is destabilized by the absence of Ltv1 (Strunk et al., 2011; Fig. 1 C). Ltv1 is required for efficient export of pre-40S ribosomes (Seiser et al., 2006; Merwin et al., 2014). Thus, Hrr25-dependent phosphorylation and release of Ltv1 must occur after nuclear export. How this presumed spatial regulation is achieved remains unknown, as there are pools of nuclear Hrr25 (Huh et al., 2003; Breker et al., 2014). It is possible that the nuclear pools of Hrr25 are not freely diffusible and instead interact with distinct structures, e.g. microtubules, which is consistent with Hrr25 having a role in spindle assembly (Petronczki et al., 2006; Rumpf et al., 2010). Alternatively, or additionally, it is possible that Hrr25-dependent Ltv1 release is slow relative to nuclear export, which is consistent with the observation that Ltv1 release is the rate-limiting step in 40S ribosome maturation (see “Kinase-mediated regulation of 40S ribosome assembly”). Finally, additional levels of regulation might be present, perhaps involving the recruitment or release of other AFs, that could regulate Hrr25 access or activity.
Affiliation: Department of Cancer Biology and Department of Chemistry, The Scripps Research Institute, Jupiter, FL 33458.