PATIENTS AND METHODS: We report two siblings of a healthy but consanguineous Malaysian family presenting with severe short stature caused by CPHD with a variable phenotype. Importantly, at the beginning the girl presented with isolated GHD, whereas the boy was hypothyroid. As the most common gene alterations responsible for CPHD are within either the PROP-1- or the POU1F1- (PIT-1)-gene these two genes were further studied.
RESULTS: Subsequent sequencing of the six exons of the POU1F1-gene allowed the identification of a new N-terminal mutation (Q4ter) in these two children. A substitution of C to T induced a change from a glutamine (CAA) to a stop codon (TAA) in exon 1 of the PIT-1 protein. Both affected children were homozygous for the mutation, whereas the mother and father were heterozygous.
CONCLUSION: We describe two children with autosomal recessive inherited CPHD caused by a new N-terminal located mutation within the PUO1F1-gene. The clinical history of these two children underline the phenotypic variability and support the fact that children with any isolated and/or combined PHD need to be closely followed as at an any time other hormonal deficiencies may occur. In addition, molecular analysis of the possible genes involved might be most helpful for the future follow-up.
RESULTS: The present report is a continuing study from our previous published transcriptomic profiling of oil palm seedlings against G. boninense. We focused on identifying differentially expressed genes (DEGs) encoding transcription factors (TFs) from the same RNA-seq data; resulting in 106 upregulated and 108 downregulated TFs being identified. The DEGs are involved in four established defense-related pathways responsible for cell wall modification, reactive oxygen species (ROS)-mediated signaling, programmed cell death (PCD) and plant innate immunity. We discovered upregulation of JUNGBRUNNEN 1 (EgJUB1) during the fungal biotrophic phase while Ethylene Responsive Factor 113 (EgERF113) demonstrated prominent upregulation when the palm switches to defense against necrotrophic phase. EgJUB1 was shown to have a binding activity to a 19 bp palindromic SNBE1 element, WNNYBTNNNNNNNAMGNHW found in the promoter region of co-expressing EgHSFC-2b. Further in silico analysis of promoter regions revealed co-expression of EgJUB1 with TFs containing SNBE1 element with single nucleotide change at either the 5th or 18th position. Meanwhile, EgERF113 binds to both GCC and DRE/CRT elements promoting plasticity in upregulating the downstream defense-related genes. Both TFs were proven to be nuclear-localized based on subcellular localization experiment using onion epidermal cells.
CONCLUSION: Our findings demonstrated unprecedented transcriptional reprogramming of specific TFs potentially to enable regulation of a specific set of genes during different infection phases of this hemibiotrophic fungal pathogen. The results propose the intricacy of oil palm defense response in orchestrating EgJUB1 during biotrophic and EgERF113 during the subsequent transition to the necrotrophic phase. Binding of EgJUB1 to SNBE motif instead of NACBS while EgERF113 to GCC-box and DRE/CRT motifs is unconventional and not normally associated with pathogen infection. Identification of these phase-specific oil palm TFs is important in designing strategies to tackle or attenuate the progress of infection.
Methods: Two complementary approaches, saturated transposon mutagenesis and spontaneous mutation induction with high concentrations of colistin and polymyxin B, were employed to select for mutations associated with resistance to polymyxins. Mutants were identified using transposon-directed insertion-site sequencing or Illumina WGS. A resistance phenotype was confirmed by MIC and further investigated using RT-PCR. Competitive growth assays were used to measure fitness cost.
Results: A transposon insertion at nucleotide 41 of the pmrB gene (EC958pmrB41-Tn5) enhanced its transcript level, resulting in a 64- and 32-fold increased MIC of colistin and polymyxin B, respectively. Three spontaneous mutations, also located within the pmrB gene, conferred resistance to both colistin and polymyxin B with a corresponding increase in transcription of the pmrCAB genes. All three mutations incurred a fitness cost in the absence of colistin and polymyxin B.
Conclusions: This study identified the pmrB gene as the main chromosomal target for induction of colistin and polymyxin B resistance in E. coli.
METHODS: We retrieved 4 previously reported SMCA, performed additional immunohistochemical and targeted next-generation sequencing (NGS). We also investigated the use of NKX3.1 as a marker for SMCA in the context of its prevalence and extent (using H-score) in a mixed cohort of retrospectively and prospectively tested head and neck lesions (n = 223) and non-neoplastic tissues (n = 66).
RESULTS: NKX3.1 positivity was confirmed in normal mucous acini as well as in mucous acinar class of lesions (5/6, mean H-score: 136.7), including mucinous adenocarcinomas (3/4), SG-IPMN (1/1), and microsecretory adenocarcinoma (MSA) (1/1). All SMCA were positive. Fluorescence in situ hybridization for SS18 rearrangements were negative in all successfully tested cases (0/3). NGS was successful in two cases (cases 3 and 4). Case 3 demonstrated a PTEN c.655C>T p.Q219* mutation and a SEC16A::NOTCH1 fusion while case 4 (clinically aggressive) showed a PTEN c.1026+1G>A p.K342 splice site variant, aTP53 c.524G>A p.R175H mutation and a higher tumor mutation burden (29 per Mb). PTEN immunohistochemical loss was confirmed in both cases and a subset of tumor cells showed strong (extreme) staining for P53 in Case 4.
CONCLUSION: Despite a partial myoepithelial phenotype, SMCA, along with mucinous adenocarcinomas/SG-IPMN and MSA, provisionally constitute a mucous acinar class of tumors based on morphology and NKX3.1 expression. Like salivary mucinous adenocarcinomas/SG-IPMN, SMCA also show alterations of the PTEN/PI3K/AKT pathway and may show progressive molecular alterations. We document the first extramammary tumor with a SEC16A::NOTCH1 fusion.