The central apparatus is an essential component of “9+2” cilia. Zhao et al. identify more than 40 new potential components of the central apparatus of Chlamydomonas. Many are conserved and will facilitate genetic screening of patients with a form of primary ciliary dyskinesia that is difficult to diagnose.

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We show that NITRATE REGULATORY GENE2 (NRG2), which we identified using forward genetics, mediates nitrate signaling in Arabidopsis thaliana. A mutation in NRG2 disrupted the induction of nitrate-responsive genes after nitrate treatment by an ammonium-independent mechanism. The nitrate content in roots was lower in the mutants than in the wild type, which may have resulted from reduced expression of NRT1.1 (also called NPF6.3, encoding a nitrate transporter/receptor) and upregulation of NRT1.8 (also called NPF7.2, encoding a xylem nitrate transporter). Genetic and molecular data suggest that NRG2 functions upstream of NRT1.1 in nitrate signaling. Furthermore, NRG2 directly interacts with the nitrate regulator NLP7 in the nucleus, but nuclear retention of NLP7 in response to nitrate is not dependent on NRG2. Transcriptomic analysis revealed that genes involved in four nitrogen-related clusters including nitrate transport and response to nitrate were differentially expressed in the nrg2 mutants. A nitrogen compound transport cluster containing some members of the NRT/ PTR family was regulated by both NRG2 and NRT1.1, while no nitrogen-related clusters showed regulation by both NRG2 and NLP7. Thus, NRG2 plays a key role in nitrate regulation in part through modulating NRT1.1 expression and may function with NLP7 via their physical interaction.

For patients with acute cholecystitis, ELC appears as safe and effective as DLC. ELC might be associated with lower hospital costs, fewer work days lost, and greater patient satisfaction.

Recent data have implicated nuclear factor B (NF-B

We report the construction of chimeric coxsackievirus B3 (CVB3) strains in which sequences of an infectious cDNA copy of a noncardiovirulent CVB3 genome were replaced by the homologous sequences from a cardiovirulent CVB3 genome to identify which of 10 predicted genetic sites determine cardiovirulence. Cardiovirulent phenotype expression was consistently linked to nucleotide 234 (U in cardiovirulent CVB3 and C in avirulent CVB3) in the 5 nontranslated region. Reconstructions of the parental noncardiovirulent CVB3 genome from chimeras restored the noncardiovirulent phenotype when tested in mice. Inoculation of severe combined immunodeficient (scid) mice with the noncardiovirulent CVB3 strain resulted in massive cardiomyocyte necrosis in all animals. Sequence analysis of viral genomes isolated from twelve scid mouse hearts showed that only nucleotide position 234 was different (a C3U transition) from that in the input parental noncardiovirulent CVB3 genome. Higher-order RNA structures predicted by two different algorithms did not demonstrate an obvious local effect caused by the C3U change at nucleotide 234. Initial studies of parental and chimeric CVB3 replication in primary cultures of fetal murine heart fibroblasts and in adult murine cardiac myocytes demonstrated that viral RNA transcriptional efficiency is approximately 10-fold lower for noncardiovirulent CVB3 than for cardiovirulent CVB3. CVB3 did not shut off protein synthesis in murine cardiac fibroblasts, nor were levels of viral protein synthesis significantly different as a function of viral phenotype. Taken together, these data support a significant role for determination of the CVB3 cardiovirulence phenotype by nucleotide 234 in the 5 nontranslated region, possibly via a transcriptional mechanism.

BackgroundSalt stress is one of the major abiotic stresses affecting plant growth and productivity. Vacuolar H+-pyrophosphatase (H+-PPase) genes play an important role in salt stress tolerance in multiple species.ResultsIn this study, the promoter from the vacuolar H+-pyrophosphatase from Thellungiella halophila (TsVP1) was cloned and compared with the AVP1 promoter from Arabidopsis thaliana. Sequence analysis indicated that these two promoters had seven similar motifs at similar positions. To determine which tissues the two promoters are active in, transgenic plants were produced with expression of the GUS reporter gene under the control of one of the promoters. In transgenic Arabidopsis with the TsVP1 promoter, the GUS reporter gene had strong activity in almost all tissues except the seeds and the activity was induced in both shoots and roots, especially in the root tips, when treated with salt stress. Such induction was not found in transgenic Arabidopsis with the AVP1 promoter. By analyzing different 5' deletion mutants of the TsVP1 promoter, an 856 bp region (-2200 to -1344) was found to contain enhancer elements that increased gene expression levels. Two AAATGA motifs, which may be the key elements for the anther specific expression profile, in the deleted TsVP1 promoters (PT2 to PT6) were also identified. A 130 bp region (-667 to -538) was finally identified as the key sequence for the salt stress response by analyzing the different mutants both with and without salt stress. GUS transient assay in tobacco leaves suggested the 130 bp region was sufficient for the salt stress response. Bioinformatic analysis also revealed that there may be novel motifs in this region that are the key elements for the salt stress responsive activity of the TsVP1 promoter.ConclusionsThe TsVP1 promoter had strong activity in almost all tissues except the seeds. In addition, its activity was induced by salt stress in leaves and roots, especially in root tips. A 130 bp region (-667 to -538) was identified as the key region for responding to salt stress.

Running title: C1a-e-c supercomplex of motile cilia Summary Fu et al. use a wild-type vs. mutant comparison and cryo-electron tomography of Chlamydomonas flagella to identify central apparatus (CA) subunits and visualize their location in the native 3D CA structure. The study provides a better understanding of the CA and how it regulates ciliary motility. Abbreviations:BCCP, biotin-carboxyl-carrier-protein; CA, central apparatus; CCD, charge-coupled device; cryo-ET, cryo-electron tomography; DMT, doublet microtubule; FAP, flagellar associated protein; IDA, inner dynein arm; MS, mass spectrometry; N-DRC, nexin-dynein regulatory complex; ODA, outer dynein arm; PCD, primary ciliary dyskinesia; psu, peripheral subunit. AbstractNearly all motile cilia contain a central apparatus (CA) composed of two connected singletmicrotubules with attached projections that play crucial roles in regulating ciliary motility.Defects in CA assembly usually result in motility-impaired or paralyzed cilia, which in humans causes disease. Despite their importance, the protein composition and functions of the CA projections are largely unknown. Here, we integrated biochemical and genetic approaches with cryo-electron tomography to compare the CA of wild type Chlamydomonas with CA mutants.We identified a large (>2 MDa) complex, the C1a-e-c supercomplex, that requires the PF16 protein for assembly and contains the CA components FAP76, FAP81, FAP92, and FAP216. We localized these subunits within the supercomplex using nanogold-labeling and show that loss of any one of them results in impaired ciliary motility. These data provide insight into the subunit organization and three-dimensional (3D) structure of the CA, which is a prerequisite for understanding the molecular mechanisms by which the CA regulates ciliary beating. Despite many biochemical and structural studies of the CA (Witman et al., 1978; protein composition, 3D organization, and functional mechanism(s) of the CA in ciliary motility are not fully understood. Our recent mass spectrometry (MS) study compared the proteomes of Chlamydomonas wild-type and mutant axonemes, and identified 44 new candidate CA proteins assigned to the C1 or the C2 microtubule (Zhao et al., 2019).However, questions about the organization, assembly, and function of the CA and its projections remain, making the CA the structurally and functionally least understood axonemal complex to date.Here we combined biochemical, genetic, and structural analyses to investigate the protein composition and molecular organization of a group of interconnected CA projections, here termed the C1a-e-c supercomplex, in wild-type and CA mutants of Chlamydomonas. Sucrose gradient sedimentation and MS revealed that several CA proteins identified in this study, FAP76, FAP81, FAP92 and FAP216, are associated with the protein PF16, previously assigned to the C1 microtubule but not to a specific projection. Chlamydomonas mutants that lacked any of these proteins showed impaired motility. Structural comparisons of flagella from wild-ty...