PilT mutations lead to simultaneous defects in competence for natural transformation and twitching motility in piliated Neisseria gonorrhoeae

Neisseria gonorrhoeae, the Gram-negative aetiological agent of gonorrhoeae, is one of many mucosal pathogens of man that expresses competence for natural transformation. Expression of this phenotype by gonococci appears to rely on the expression of type IV pili (Tfp), but the mechanistic basis for this relationship remains unknown. During studies of gonococcal pilus biogenesis, a homologue of the PilT family of proteins, required for Tfp-dependent twitching motility in Pseudomonas aeruginosa and social gliding motility in Myxococcus xanthus, was discovered. Like the findings in these other species, we show here that gonococcal PilT mutants constructed in vitro no longer display twitching motility. In addition, we demonstrate that they have concurrently lost the ability to undergo natural transformation, despite the expression of structurally and morphologically normal Tpf. These results were confirmed by the findings that two classes of spontaneous mutants that failed to express twitching motility and transformability carried mutations in PilT. Piliated PilT mutants and a panel of pilus assembly mutants were found to be deficient in sequence-specific DNA uptake into the cell, the earliest demonstrable step in neisserial competence. The PilT-deficient strains represent the first genetically defined mutants that are defective in DNA uptake but retain Tfp expression.     

Induction of forward mutations at the thymidine kinase locus of mouse lymphoma cells: evidence for electrophilic and non-electrophilic mechanisms

A database of 209 chemicals tested for induction of forward mutations at the heterozygous thymidine kinase (TK +/-) locus in L5178Y mouse lymphoma cells was analyzed for structure-activity relationships using the MultiCASE expert system. Consistent with evidence of several contributing biological mechanisms, these studies suggest that such mutations may occur by more than one mechanism. As might be expected, there was evidence for a component involving direct electrophilic attack on the cellular DNA, in a manner previously established as causative in the induction of mutations in Salmonella. In addition, however, there was also strong evidence for another mechanism or mechanisms involving chromosome missegregation, cellular toxicity or an alternate site of action, such as the microtubules.     

Refining Genotypes and Phenotypes in KCNA2-Related Neurological Disorders

Pathogenic variants in KCNA2, encoding for the voltage-gated potassium channel K_v1.2, have been identified as the cause for an evolving spectrum of neurological disorders. Affected individuals show early-onset developmental and epileptic encephalopathy, intellectual disability, and movement disorders resulting from cerebellar dysfunction. In addition, individuals with a milder course of epilepsy, complicated hereditary spastic paraplegia, and episodic ataxia have been reported. By analyzing phenotypic, functional, and genetic data from published reports and novel cases, we refine and further delineate phenotypic as well as functional subgroups of KCNA2-associated disorders. Carriers of variants, leading to complex and mixed channel dysfunction that are associated with a gain- and loss-of-potassium conductance, more often show early developmental abnormalities and an earlier onset of epilepsy compared to individuals with variants resulting in loss- or gain-of-function. We describe seven additional individuals harboring three known and the novel KCNA2 variants p.(Pro407Ala) and p.(Tyr417Cys). The location of variants reported here highlights the importance of the proline(405)–valine(406)–proline(407) (PVP) motif in transmembrane domain S6 as a mutational hotspot. A novel case of self-limited infantile seizures suggests a continuous clinical spectrum of KCNA2-related disorders. Our study provides further insights into the clinical spectrum, genotype–phenotype correlation, variability, and predicted functional impact of KCNA2 variants.     

Solubility of Si3N4 in Liquid SiO2

The nitrogen solubility in the SiO₂-rich liquid in the metastable binary SiO₂-Si₃N₄ system has been determined by analytical TEM to be 1%–4% of N/(O + N) at 1973–2223 K. Analysis of the near edge structure of the electron energy loss peak indicates that nitrogen is incorporated into the silicate network rather than being present as molecular N₂. A regular solution model with a positive enthalpy of mixing for the liquid was used to match the data for the metastable solubility of N in the presence of crystalline Si₃N₄ and to adjust the computed phase diagram. The solubility of Si₃N₄ in fused SiO₂ is far less than reported in liquid silicates also containing Al, Mg, and/or Y. Apparently, these cations act as modifiers that break anion bridges in the silicate network and, thereby, allow further incorporation of Si₃N₄ without prohibitive amounts of network cross-linking. Finally, indications emerged regarding the diffuse nature of the Si₃N₄-SiO₂ interface that leads to amorphous regions of higher N content.