RNA modifications represent a novel layer of regulation of gene expression. Functional experiments revealed that N 6-methyladenosine (m 6 A) on messenger RNA (mRNA) plays critical roles in cell fate determination and development. m 6 A mark also resides in the decoding center of 18S ribosomal RNA (rRNA); however, the biological function of m 6 A on 18S rRNA is still poorly understood. Here, we report that methyltransferase-like 5 (METTL5) methylates 18S rRNA both in vivo and in vitro, which is consistent with previous reports. Deletion of Mettl5 causes a dramatic differentiation defect in mouse embryonic stem cells (mESCs). Mechanistically, the m 6 A deposited by METTL5 is involved in regulating the efficient translation of F-box and WD repeat domain-containing 7 (FBXW7), a key regulator of cell differentiation. Deficiency of METTL5 reduces FBXW7 levels and leads to the accumulation of its substrate c-MYC, thereby delaying the onset of mESC differentiation. Our study uncovers an important role of METTL5-mediated 18S m 6 A in mESC differentiation through translation regulation and provides new insight into the functional significance of rRNA m 6 A.

Motile cilia and flagellar defects can result in primary ciliary dyskinesia (PCD), which is a multisystemic genetic disorder that affects roughly 1:10000 individuals. The nexin-dynein regulatory complex (N-DRC) links neighboring doublet microtubules within flagella, serving as a central regulatory hub for motility in Chlamydomonas. Herein, we identified two homozygous DRC1 variants in human patients that were associated with multiple morphological abnormalities of the sperm flagella (MMAF) and male infertility. Drc1−/−, Drc1R554X/R554X, and Drc1W244X/W244X mice on the C57BL/6 background suffered from prepubertal mortality. However, when the ICR background was introduced, some of these mice were able to survive and recapitulate the MMAF phenotypes detected in human patients. By analyzing these animals, we determined that DRC1 is an essential regulator of N-DRC assembly in cilia and flagella. When DRC1 is absent, this results in the shortening of cilia and consequent impairment of their motility. Damage associated with DRC1 deficiency in sperm flagella was more pronounced than in cilia, as manifested by complete axoneme structural disorder in addition to the loss of the DRC structure. Together, these findings suggest that DRC1 is required for the structural stability of flagella but not cilia, emphasizing the key role of this protein in mammalian species.

Cilia and flagella are ancient structures that achieve controlled motor functions through the coordinated interaction based on microtubules, and some attached projections. Radial spokes (RSs) facilitate the beating motion of these organelles by mediating signal transduction between dyneins and a central pair (CP) of singlet microtubules. RS complex isolation from Chlamydomonas axonemes enabled the detection of 23 radial spoke proteins (RSP1-23), with the roles of some radial spoke proteins remained unknown. Recently, RSP15 has been reported to be located to the stalk of RS2, but its homolog in mammals has not been explored. Herein, we show that Lrrc23 is an evolutionarily conserved testis-enriched gene encoding an RSP15 homolog in mice. We found that LRRC23 localizes to the RS complex within murine sperm flagella and interacts with RSPH3A/B. The knockout of Lrrc23 resulted in male infertility due to RS disorganization and impaired motility in murine spermatozoa, whereas the ciliary beating was unaffected significantly. These data indicate that LRRC23 is a key regulator underpinning the integrity of RS complex within the flagella of mammalian spermatozoa, whereas it is dispensable in cilia.

Progressive cerebral infarction (PCI) is associated with high rates of mortality and disability. Many studies have shown that Dl-3n-butylphthalide (NBP) is effective against acute ischemic stroke. The administration of NBP can result in an increased number of capillaries in the ischemic region, promote the establishment of collateral circulation, protect the mitochondria, and narrow the infarction area, among other effects. In the present study, we evaluated the efficacy and safety of NBP for the treatment of PCI.Between March 2008 and May 2012, we performed a randomized, double-blind placebo-controlled study including 304 inpatients with PCI. These patients were randomly assigned to the test (152 cases) and control groups (152 cases). The test group received 200 mg of NBP soft capsules orally, 15 minutes before each meal, 3 times daily. The control group received 200 mg of placebo soft capsules orally, 15 minutes before each meal, 3 times daily. Treatment was administered during 21 days. The National Institute of Health Stroke Scale (NIHSS) score was assessed before the treatment and on days 7, 14, 21, and 30 after treatment. The Barthel index (BI) was assessed on the same days and on day 90.In the test group, the NIHSS scores on days 7, 14, 21, and 30 were 14.75 ± 4.85, 11.62 ± 3.49, 8.87 ± 5.17, and 6.38 ± 4.93, respectively. In the control group, they were 16.08 ± 3.76, 13.28 ± 5.02, 11.05 ± 4.25, and 8.43 ± 5.41 (P < .05), respectively. The BI on days 7, 14, 21, 30, and 90 were 51.57 ± 15.11, 61.21 ± 16.39, 70.48 ± 18.21, 76.41 ± 19.02, and 81.10 ± 15.52 for the test group and 46.79 ± 18.42, 55.93 ± 19.12, 64.84 ± 17.67, 70.65 ± 18.54, and 76.54 ± 17.05 for the control group (P < .05), respectively. Adverse events were elevation of alanine aminotransferase and aspartate aminotransferase (P > .05).NBP was useful to improve the outcome of patients with PCI and decreased their disability for activities of daily living. NBP was an efficacious and safe treatment for PCI.

Defects in the structure or motility of cilia and flagella may lead to severe diseases such as primary ciliary dyskinesia (PCD), a multisystemic disorder with heterogeneous manifestations affecting primarily respiratory and reproductive functions. We report that CFAP61 is a conserved component of the Calmodulin and radial Spoke associated Complex (CSC) of cilia. We find that a CFAP61 splice variant, c.143+5G&gt;A, causes exon skipping/ intron retention in human, inducing a multiple morphological abnormalities of the flagella (MMAF) phenotype. We generated Cfap61 knockout mice that recapitulate the infertility phenotype of the human CFAP61 mutation, but without other symptoms usually observed in PCD. We find that CFAP61 interacts with the CSC, radial spoke (RS) stalk and head. During early stages of Cfap61−/- spermatid development, the assembly of RS components is impaired. With the progress of spermiogenesis, the axoneme in Cfap61−/- cells becomes unstable and scatters, and the distribution of intraflagellar transport proteins is disrupted. This study reveals an organ specific mechanism of axoneme stabilization that is related to male infertility.