Mapping of a gene determining tuberous sclerosis to human chromosome 11q14-11q23

Tuberous sclerosis (TSC) is a dominantly inherited disorder characterized by hamartomas and hamartias in one or more organs, most often in skin, brain, and kidneys. Analysis of the basic genetic defect in tuberous sclerosis would be greatly expedited by definitive determination of the chromosomal location of the TSC gene or genes. We have carried out genetic linkage studies in 15 TSC families, using 34 polymorphic markers including protein markers and DNA markers. Pairwise lod scores were calculated using LIPED, and multipoint analyses were carried out using MENDEL. In the pairwise linkage analysis, using a penetrance value of 90%, a significant positive lod score was obtained with MCT128.1 (D11S144), 11q22-11q23, Zmax 3.26 at theta = 0.08. The tyrosinase probe TYR (11q14-11q22) gave a maximum lod score of 2.88 at theta = 0. In the multipoint analyses the most likely order is (TYR,TSC)-MCT128.1-HHH172. Homogeneity analysis was carried out using the USERM9 subprogram of MENDEL, which conducts the admixture test of C. Smith (1963, Ann. Hum. Genet. 27: 175-182). This test provided no evidence for genetic heterogeneity (that is, non-11-linked families) in this data set.     

Proposed virulence factors among coagulase-negative staphylococci isolated from two healthy populations

Proposed virulence factors, including multiple antibiotic resistance and slime-mediated adherence, among coagulase-negative staphylococci colonizing healthy individuals from two different study populations were examined. Resistance to methicillin was more commonly seen than initially anticipated. In addition, adherence characteristics, as quantitated by a microtiter plate method, were very similar to those of strains of coagulase-negative staphylococci causing nosocomial infections.     

Natural occurrence of Fusarium mycotoxins in field samples from the 1992 Wisconsin corn crop.

Analysis of 98 moldy corn samples collected in Wisconsin between November 1992 and January 1993 for Fusarium toxins by various immunochemical assays revealed overall average mycotoxin concentrations of 305.6, 237.7, and 904.3 ng/g for type A trichothecenes (TCTCs), deoxynivalenol (DON)-related type B TCTCs (total DON), and zearalenone (ZE), respectively. A small portion (5.1%) of the samples was found to be contaminated with high levels ( > 1 microgram/g) of type A TCTCs and total DON during the whole survey. Over 40% of the samples had 100 to 1,000 ng of total DON per g, while 17% of the samples had the same levels of type A TCTCs. The analytical data were consistent with those from mycological examinations for the samples in which various toxic Fusarium spp., including F. sporotrichioides, F. poae, and F. graminearum, were found. The samples received in November 1992 had relatively low concentrations of toxin; the average levels of type A TCTCs and total DON were 9.9 and 79 ng/g, respectively. The toxin concentrations became progressively higher in the samples received in December. The average levels for the type A TCTCs and total DON increased to 920 and 335 ng/g, respectively. However, the levels of ZE were higher in the samples collected earlier. The average levels for samples collected in November and late December were 1,195 and 242 ng/g, respectively. Analysis of selected samples by high-performance liquid chromatography monitoring with an enzyme-linked immunosorbent assay revealed that T-2 toxin, HT-2 toxin, diacetoxyscirpenol, neosolaniol, and T-2 tetraol (T-2-4ol) were common in these samples. Statistical analysis revealed a weak correlation between the levels of total type A TCTCs and total DON in the samples (r = 0.18, P = 0.09), but a strong correlation between the levels of ZE and total type B TCTCs (r = 0.75, P < 0.0001) was found. The mycotoxin levels of total type A TCTCs, total DON-related type B TCTCs, and ZE in the cobs (5.2, 3.9, and 21 micrograms/g, respectively) were considerably higher than those in the kernels (1.0, 0.5, and 0.5 microgram/g, respectively). The type A toxin levels increased from a range of 14 to 35 ng/g to a range of 110 to 538 ng/g after the moldy corn samples were held at 5 degrees C for 8 days in the laboratory.     

The growth and nutrient uptake of winter wheat

Summary Measurements with an ion selective electrode under winter wheat and in adjacent fallow soil, from April to July 1976, showed that nitrate concentrations were high in the 0–25 cm zone and correspondingly lower at 50 cm, because of the extreme drying conditions. Maximum differences in nitrate concentrations between cropped and fallow soil occurred at Feekes' stages 6, 10, and 11.1 indicating periods of maximum uptake by the crop (cf Ref.⁴).Dry matter weight of wheat, sampled biweekly, was maximum 15 days before maturity. The foliage senesced and lost weight from Feekes' stage 10.1 onwards. Nutrient concentrations in the foliage decreased from Feekes' stage 4, but N, P and Mg concentrations in the ears increased during Feekes' stage 11. N, P and Mg accumulated in the ears at the expense of the foliage during stage 11, maximum uptake occurring at stages 11.3, 11.1 and 11.2 respectively. In contrast, K and Ca uptake ceased at stages 10.1 and 10.5 respectively and then both were lost from the foliage in heavy showers.Rates of N uptake and soil nitrate depletion correlated significantly, enabling N uptake to be deduced approximately from thesein situ soil nitrate measurements.     

Study of the structure of troponin-T by measuring the relative reactivities of lysines with acetic anhydride

The structure of troponin-T has been studied by measuring the relative reactivity of lysines with acetic anhydride using a competitive labeling method. Troponin-T was acetylated free and complexed with -I and -C in the native state with [³H]acetic anhydride, purified, and then combined with ¹⁴[C]troponin-T that had been acetylated in 6 m-guanidine · HCl. Peptides containing labeled lysines were isolated following chymotryptic and tryptic digestion and identified in the published sequence. The ${}^3\text{H}{}^{14}\text{C}$ ratio of these peptides was used as a measure of relative accessibility of the lysines. Troponin-T contains 39 lysines; we have identified 35 of these in 22 different peptides. The region of troponin-T influenced by binding to the other troponin components is extensive and includes the C-terminal half of the molecule as well as some residues in the N-terminal half. The lysines showing the greatest change in reactivity are concentrated between residues 114 to 223. The reactivities of the troponin-T lysines labeled in native troponin were not significantly influenced by the binding of calcium to the calcium-specific binding sites of troponin-C. A model for the structure of troponin-T is proposed based on the present and previous studies.     

Application of Multiplexed Kinase Inhibitor Beads to Study Kinome Adaptations in Drug-Resistant Leukemia

Protein kinases play key roles in oncogenic signaling and are a major focus in the development of targeted cancer therapies. Imatinib, a BCR-Abl tyrosine kinase inhibitor, is a successful front-line treatment for chronic myelogenous leukemia (CML). However, resistance to imatinib may be acquired by BCR-Abl mutations or hyperactivation of Src family kinases such as Lyn. We have used multiplexed kinase inhibitor beads (MIBs) and quantitative mass spectrometry (MS) to compare kinase expression and activity in an imatinib-resistant (MYL-R) and -sensitive (MYL) cell model of CML. Using MIB/MS, expression and activity changes of over 150 kinases were quantitatively measured from various protein kinase families. Statistical analysis of experimental replicates assigned significance to 35 of these kinases, referred to as the MYL-R kinome profile. MIB/MS and immunoblotting confirmed the over-expression and activation of Lyn in MYL-R cells and identified additional kinases with increased (MEK, ERK, IKKα, PKCβ, NEK9) or decreased (Abl, Kit, JNK, ATM, Yes) abundance or activity. Inhibiting Lyn with dasatinib or by shRNA-mediated knockdown reduced the phosphorylation of MEK and IKKα. Because MYL-R cells showed elevated NF-κB signaling relative to MYL cells, as demonstrated by increased IκBα and IL-6 mRNA expression, we tested the effects of an IKK inhibitor (BAY 65-1942). MIB/MS and immunoblotting revealed that BAY 65-1942 increased MEK/ERK signaling and that this increase was prevented by co-treatment with a MEK inhibitor (AZD6244). Furthermore, the combined inhibition of MEK and IKKα resulted in reduced IL-6 mRNA expression, synergistic loss of cell viability and increased apoptosis. Thus, MIB/MS analysis identified MEK and IKKα as important downstream targets of Lyn, suggesting that co-targeting these kinases may provide a unique strategy to inhibit Lyn-dependent imatinib-resistant CML. These results demonstrate the utility of MIB/MS as a tool to identify dysregulated kinases and to interrogate kinome dynamics as cells respond to targeted kinase inhibition.     

Melanoma central nervous system metastases: An update to approaches,challenges, and opportunities

In February 2018, the Melanoma Research Foundation and the Moffitt Cancer Center hosted the Second Summit on Melanoma Central Nervous System (CNS) Metastases in Tampa, Florida. In this white paper, we outline the current status of basic science, translational, and clinical research into melanoma brain metastasis development and therapeutic management. We further outline the important challenges that remain for the field and the critical barriers that need to be overcome for continued progress to be made in this clinically difficult area.     

Guanine-rich telomeric sequences stimulate DNA polymerase activity in vitro

Guanine-rich oligonucleotides and short telomeric DNA sequences can self-associate into G-quartet stabilized complexes. We discovered that this self-association can occur in sequencing reactions and that higher-order structures stimulate DNA polymerase to synthesize extended DNA strands. Base analogues were used to identify Hoogsteen base pairings as stabilizing forces in these stimulatory DNA structures. Scanning force microscopy confirmed that quartet-DNA was formed from these oligomers and that these extended, four-stranded structures could be bound by DNA polymerase. Since guanine quartet-stabilized structures are proposed to exist in vivo, such structures may stimulate DNA polymerization in vivo.     

Novel pyrazolopyrimidine derivatives as GSK-3 inhibitors

A series of [1-aryl-1H-pyrazolo[3,4-d]pyrimidin-4-yl]arylhydrazones were discovered as novel inhibitors glycogen synthase kinase-3 (GSK-3). Based on initial modeling a detailed SAR was constructed. Modification of the interior binding aryl ring (Ar(1)) determined this to be a tight binding region with little room for modification. As predicted from the model, a large variety of modifications could be incorporated into the hydrazone aryl ring. This work led to GSK-3 inhibitors in the low nano-molar range.