Circumventing glucocorticoid-mediated hyperinsulinemia via the activation of PPARα

Comment on: PPARα blocks glucocorticoid receptor α-mediated transactivation but cooperates with the activated glucocorticoid receptor alpha for transrepression on NFκB. Bougarne N, Paumelle R, Caron S, Hennuyer N, Mansouri R, Gervois P, Staels B, Haegeman G, De Bosscher K. Proc Natl Acad Sci USA 2009; 106:7397-402. PMID: 19376972     

Consistencies of individual DNA base-amino acid interactions in structures and sequences.

Amino acid-amino acid interaction energies have been derived from crystal structure data for a number of years. Here is reported the first derivation of normalized relative interaction from binding data for each of the four bases interacting with a specific amino acid, utilizing data from combinatorial multiplex DNA binding of zinc finger domains [Desjarlais, J. R. and Berg, J. M. (1994) Proc. Natl. Acad. Sci. USA, 91, 11099-11103]. The five strongest interactions are observed for lysine-guanine, lysine-thymine, arginine-guanine, aspartic acid-cytosine and asparagine-adenine. These rankings for interactions with the four bases appear to be related to base-amino acid partial charges. Also, similar normalized relative interaction energies are derived by using DNA binding data for Cro and lambda repressors and the R2R3 c-Myb protein domain [Takeda, Y., Sarai, A. and Rivera, V. M. (1989) Proc. Natl. Acad. Sci. USA, 86, 439-443; Sarai, A. and Takeda, Y. (1989) Proc. Natl. Acad. Sci. USA, 86, 6513-6517; Ogata, K. et al. (1995) submitted]. These energies correlate well with the combinatorial multiplex energies, and the strongest cases are similar between the two sets. They also correlate well with similar relative interaction energies derived directly from frequencies of bases in the bacteriophage lambda operator sequences. These results suggest that such potentials are general and that extensive combinatorial binding studies can be used to derive potential energies for DNA-protein interactions.     

Chromosomal locations of members of a family of novel endogenous human retroviral genomes.

Human cellular DNA contains two distinguishable families of retroviral related sequences. One family shares extensive nucleotide sequence homology with infectious mammalian type C retroviral genomes (T. I. Bonner, C. O'Connell, and M. Cohen, Proc. Natl. Acad. Sci. USA 79:4709-4713, 1982; M. A. Martin, T. Bryan, S. Rasheed, and A. S. Khan, Proc. Natl. Acad. Sci. USA 78:4892-4896, 1981). The other family contains major regions of homology with the pol genes of infectious type A and B and avian type C and D retroviral genomes (R. Callahan, W. Drohan, S. Tronick, and J. Schlom, Proc. Natl. Acad. Sci. USA 79:5503-5507, 1982; I. M. Chiu, R. Callahan, S. R. Tronick, J. Schlom, and S. A. Aaronson, Science 223:364-370, 1984). Analysis of the human recombinant clone HLM-2 has shown that the pol gene in the latter family is located within an endogenous proviral genome (R. Callahan, I. M. Chiu, J. F. H. Wong, S. R. Tronick, B. A. Roe, S. A. Aaronson, and J. Schlom, Science 228:1208-1211, 1985). We show that the proviral genome in HLM-2 and the related recombinant clone HLM-25 are located, respectively, on human chromosomes 1 and 5. Other related proviral genomes are located on chromosomes 7, 8, 11, 14, and 17.     

The chimeric VirA-tar receptor protein is locked into a highly responsive state.

The wild-type VirA protein is known to be responsive not only to phenolic compounds but also to sugars via the ChvE protein (G. A. Cangelosi, R. G. Ankenbauer, and E. W. Nester, Proc. Natl. Acad. Sci. USA 87:6708-6712, 1990, and N. Shimoda, A. Toyoda-Yamamoto, J. Nagamine, S. Usami, M. Katayama, Y. Sakagami, and Y. Machida, Proc. Natl. Acad. Sci. USA 87:6684-6688, 1990). It is shown here that the mutant VirA(Ser-44, Arg-45) protein and the chimeric VirA-Tar protein are no longer responsive to sugars and the ChvE protein. However, whereas the chimeric VirA-Tar protein was found to be locked in a highly responsive state, the VirA(Ser-44, Arg-45) mutant protein appeared to be locked in a low responsive state. This difference turned out to be important for tumorigenicity of the host strains in virulence assays on Kalanchoë daigremontiana.     

DNA knots and DNA supercoiling

Comment on: Witz G, et al. Proc Natl Acad Sci USA 2011; 108:3608-11.     

Cytoplasmic sequestration of wild-type p53 protein impairs the G1 checkpoint after DNA damage.

Wild-type p53 protein is abnormally sequestered in the cytoplasm of a subset of primary human tumors including neuroblastomas (NB) (U. M. Moll, M. LaQuaglia, J. Benard, and G. Riou, Proc. Natl. Acad. Sci. USA 92:4407-4411, 1995; U. M. Moll, G. Riou, and A. J. Levine, Proc. Natl. Acad. Sci.USA 89:7262-7266, 1992). This may represent a nonmutational mechanism for abrogating p53 tumor suppressor function. To test this hypothesis, we established the first available in vitro model that accurately reflects the wild-type p53 sequestration found in NB tumors. We characterized a series of human NB cell lines that overexpress wild-type p53 and show that p53 is preferentially localized to discrete cytoplasmic structures, with no detectable nuclear p53. These cell lines, when challenged with a variety of DNA strand-breaking agents, all exhibit impaired p53-mediated G1 arrest. Induction analysis of p53 and p53-responsive genes show that this impairment is due to suppression of nuclear p53 accumulation. Thus, this naturally occurring translocation defect compromises the suppressor function of p53 and likely plays a role in the tumorigenesis of these tumors previously thought to be unaffected by p53 alterations.     

A natural oocyte component required for the reprogramming of somatic cell nuclei

Comment on: Jullien J, et al. Proc Natl Acad Sci USA 2010; 107:5483-8.     

Immunological and structural homology between human T-cell leukemia virus type I envelope glycoprotein and a region of human interleukin-2 implicated in binding the beta receptor.

The N-terminal segment of human interleukin-2 (hIL-2) appears to mediate binding of the beta hIL-2 receptor (R. Robb, C. Rusk, J. Yodoi, and W. Greene, Proc. Natl. Acad. Sci. USA 84:2002-2006, 1987). An affinity-purified antibody prepared against this peptide segment (p81) is shown here to cross-react with a homologous region of the human T-cell leukemia virus type I (HTLV-I) envelope glycoprotein, raising the interesting possibility that the envelope glycoprotein of HTLV-I can interact with the beta hIL-2 receptor.     

Enhanced survival of pluripotent stem cells under stressful conditions

Comment on: Ardehali R, et al. Proc Natl Acad Sci USA. 2011; 108:3282-7.     

ras gene Amplification and malignant transformation.

Morphologic transformation of NIH 3T3 mouse cells occurs upon transfection of these cells with large amounts (greater than or equal to 10 micrograms) of recombinant DNA molecules carrying the normal human H-ras-1 proto-oncogene. We provide experimental evidence indicating that transformation of these NIH 3T3 cells results from the combined effect of multiple copies of the H-ras-1 proto-oncogene rather than from spontaneous mutation of one of the transfected H-ras-1 clones (E. Santos, E.P. Reddy, S. Pulciani, R.J. Feldman, and M. Barbacid, Proc. Natl. Acad. Sci. USA 80:4679-4683, 1983). Levels of H-ras-1 RNA and p21 expression are highly elevated in the NIH 3T3 transformants, and in those cases examined, these levels correlate with the malignant properties of these cells. We have also investigated the presence of amplified ras genes in a variety of human carcinomas. In 75 tumor biopsies, we found amplification of the human K-ras-2 locus in one carcinoma of the lung. These results indicate that ras gene amplification is an alternative pathway by which ras genes may participate in the development of human neoplasia.     

Failure to transmit disease from gray tremor mutant mice.

Mice homozygous for mutant alleles at the gray tremor (gt) locus develop a marked non-intention tremor beginning at 8 days of age. Most homozygous mice die by 3 months. Homozygotes exhibit intense vacuolation of the central nervous system gray matter and vacuolation and hypomyelination of some white matter tracts. Based on neuropathological similarities with scrapie, other investigators inoculated wild-type mice with gray tremor brain homogenates to test the hypothesis of transmissibility. Published reports indicated that spongiform encephalopathy (R. L. Sidman, H. C. Kinney, and H. O. Sweet, Proc. Natl. Acad. Sci. USA 82:253-257, 1985) and disease, including hind limb paralysis in NFS mice (P. M. Hoffman, R. G. Rohwer, C. MacAuley, J. A. Bilello, J. W. Hartley, and H. C. Morse III, Proc. Natl. Acad. Sci. USA 84:3866-3870, 1987), were transmitted by inoculation of gt/gt brain homogenates. In our hands, however, no NFS/NCr animals inoculated intracerebrally with gt/gt or +/+ brain preparations showed any signs of disease or pathological changes in the brain. Positive transmission by other investigators may reflect the microbiological status of their donor or recipient mice.     

Delineating oncogene/tumor suppressor interactions in human mammary epithelial cells

Comment on: Cipriano R, et al. Proc Natl Acad Sci USA 2011; 108:8668-73.     

Percolation model of ionic channel dynamics.

The nonexponential closed-time distributions observed for ionic channels have been explained recently by quasi-one-dimensional models of structural diffusion (Millhauser, G. L., E. E. Salpeter, and R. E. Oswald. 1988. Proc. Natl. Acad. Sci. USA. 85: 1503-1507; Condat, C. A., and J. J?ckle. 1989. Biophys. J. 55: 915-925; Levitt, D. G. 1989. Biophys. J. 55: 489-498). We generalize this treatment by allowing for more complex trajectories using percolation theory. We assume that the gating transition depends on marginally connected conformational states leading to the observed spread in time scales.     

Intracellular transport and sorting of mutant human proinsulins that fail to form hexamers

Human proinsulin and insulin oligomerize to form dimers and hexamers. It has been suggested that the ability of prohormones to self associate and form aggregates may be responsible for the sorting process at the trans-Golgi. To examine whether insulin oligomerization is required for proper sorting into regulated storage granules, we have constructed point mutations in human insulin B chain that have been previously shown to prevent formation of insulin hexamers (Brange, J., U. Ribel, J. F. Hansen, G. Dodson, M. T. Hansen, S. Havelund, S. G. Melberg, F. Norris, K. Norris, L. Snel, A. R. Sorensen, and H. O. Voight. 1988. Nature [Lond.]. 333:679-682). One mutant (B10His----Asp) allows formation of dimers but not hexamers and the other (B9Ser----Asp) prevents formation of both dimers and hexamers. The mutants were transfected into the mouse pituitary AtT-20 cells, and their ability to be sorted into regulated secretory granules was compared to wild-type insulin. We found that while B10His----Asp is sorted somewhat less efficiently than wild-type insulin as reported previously (Carroll, R. J., R. E. Hammer, S. J. Chan, H. H. Swift, A. H. Rubenstein, and D. F. Steiner. 1988. Proc. Natl. Acad. Sci. USA. 85:8943-8947; Gross, D. J., P. A. Halban, C. R. Kahn, G. C. Weir, and L. Villa-Kumaroff. 1989. Proc. Natl. Acad. Sci. USA. 86:4107-4111). B9Ser----Asp is targeted to granules as efficiently as wild-type insulin. These results indicate that self association of proinsulin into hexamers is not required for its targeting to the regulated secretory pathway.     

c[D-pro-Pro-D-pro-N-methyl-Ala] adopts a rigid conformation that serves as a scaffold to mimic reverse-turns

Naturally occurring cyclic tetrapeptides (CTPs) such as tentoxin (Halloin et al., Plant Physiol 1970, 45, 310-314; Saad, Phytopathology 1970, 60, 415-418), ampicidin (Darkin-Rattray, Proc Natl Acad Sci USA 1996, 93, 13143-13147), HC-toxin (Walton, Proc Natl Acad Sci USA 1987, 84, 8444-8447), and trapoxin (Yoshida and Sugita, Jpn J Cancer Res 1992, 83, 324-328; Itazaki et al., J Antibiot (Tokyo) 1990, 43, 1524-1532) have a wide range of biological activity and potential use ranging from herbicides (Walton, Proc Natl Acad Sci USA 1987, 84, 8444-8447; Judson, J Agric Food Chem 1987, 35, 451-456) to therapeutics (Loiseau, Biopolymers 2003, 69, 363-385) for malaria (Darkin-Rattray, Proc Natl Acad Sci USA 1996, 93, 13143-13147) and cancer (Yoshida and Sugita, Jpn J Cancer Res 1992, 83, 324-328). To elucidate scaffolds that have few low-energy conformations and could serve as semirigid reverse-turn mimetics, the flexibility of CTPs was determined computationally. Four analogs of cyclic tetraproline c[Pro-pro-Pro-pro] with alternating L- and D-prolines, namely c[pro-Pro-pro-NMe-Ala], c[pip-Pro-pip-Pro], c[pro-Pip-pro-Pro], and c[Ala-Pro-pip-Pro] were synthesized and characterized by NOESY NMR. Both molecular mechanics and Density Functional Theory quantum calculations found these head-to-tail CTPs to be constrained to one or two relatively stable conformations. NMR structures, while not always yielding the same lowest energy conformation as expected by in silico predictions, confirmed only one or two highly populated solution conformations for all four peptides examined. c[pro-Pro-pro-NMe-Ala] was shown to have a single all trans-amide bond conformation from both in silico predictions and NMR characterization, and to be a reverse-turn mimetic by overlapping four Calpha-Cbeta bonds with those for approximately 6.5% (Tran, J Comput Aided Mol Des 2005, 19, 551-566) of reverse-turns in the Protein Data Bank PDB with a RMSD of 0.57 A.     

Effect of colchicine on internalization of prolactin in female rat liver: an in vivo radioautographic study

Binding and internalization of 125I-ovine prolactin into hepatocytes of female rats was visualized by the in vivo radioautographic method (Bergeron, J. J. M., G. Levine, R. Sikstrom, D. O'Shaughnessey, B. Kopriwa, N. J. Nadler, and B. I. Posner, 1977, Proc. Natl. Acad. Sci. USA, 745:051-5055). Receptor-mediated internalization of label was observed into lipoprotein-filled vesicles in the Golgi/bile canalicular region of the hepatocyte. Colchicine treatment had no effect on the internalization of label into the lipoprotein-filled vesicles. However, the location of the radio-labeled lipoprotein-filled vesicles was altered from the Golgi/bile canalicular region to subsinusoidal. Radioactive content of hepatocytes decreased as a function of time after injection of 125I-prolactin; however, colchicine treatment markedly retarded this loss of label. Subcellular fractionation experiments indicated that colchicine treatment led to decreased levels of 125I-prolactin accumulation in microsomes but augmented the accumulation of label in the L fraction. It is concluded that in normal female rats prolactin is internalized into lipoprotein-filled vesicles in the Golgi region before degradation of the hormone. Colchicine treatment accumulates labeled lipoprotein-containing vesicles in a subsinusoidal region and retards hormone catabolism. The labeled vesicles observed after colchicine treatment may correspond to the unique vesicles previously observed in the L fraction and found to be enriched in prolactin receptors (Khan, M. N., B. I. Posner, A. K. Verma, R. J. Khan, and J. J. M. Bergeron, 1981, Proc. Natl. Acad. Sci. USA, 78:4980-4981).     

The actin-based nanomachine at the leading edge of migrating cells.

Two fundamental parameters of the highly dynamic, ultrathin lamellipodia of migrating fibroblasts have been determined-its thickness in living cells (176 +/- 14 nm), by standing-wave fluorescence microscopy, and its F-actin density (1580 +/- 613 microm of F-actin/microm(3)), via image-based photometry. In combination with data from previous studies, we have computed the density of growing actin filament ends at the lamellipodium margin (241 +/- 100/microm) and the maximum force (1.86 +/- 0.83 nN/microm) and pressure (10.5 +/- 4.8 kPa) obtainable via actin assembly. We have used cell deformability measurements (. J. Cell Sci. 44:187-200;. Proc. Natl. Acad. Sci. USA. 79:5327-5331) and an estimate of the force required to stall the polymerization of a single filament (. Proc. Natl. Acad. Sci. USA. 78:5613-5617;. Biophys. J. 65:316-324) to argue that actin assembly alone could drive lamellipodial extension directly.