The capsids of spherical viruses may contain from tens to hundreds of copies of the capsid protein(s). Despite their complexity, these particles assemble rapidly and with high fidelity. Subunit and capsid represent unique end states. However, the number of intermediate states in these reactions can be enormous-a situation analogous to the protein folding problem. Approaches to accurately model capsid assembly are still in their infancy. In this paper, we describe a sailshaped reaction landscape, defined by the number of subunits in each species, the predicted prevalence of each species, and species stability. Prevalence can be calculated from the probability of synthesis of a given intermediate and correlates well with the appearance of intermediates in kinetics simulations. In these landscapes, we find that only those intermediates along the leading edge make a significant contribution to assembly. Although the total number of intermediates grows exponentially with capsid size, the number of leading-edge intermediates grows at a much slower rate. This result suggests that only a minute fraction of intermediates needs to be considered when describing capsid assembly.Keywords: capsid assembly; virus assembly; protein polymerization; protein folding; energy landscape Supplemental material: see www.proteinscience.orgComplicated reactions, such as protein folding, are often described in terms of energy landscapes (Bryngelson et al. 1995;Wolynes 1996;Brooks et al. 1998;Chan and Dill 1998;Onuchic et al. 1998;Dobson and Karplus 1999;Dinner et al. 2000). This representation of the reaction allows a quantitative description of the multiplicity of intermediates, the many paths that can be followed to local and global minima, and the different kinetic barriers that each path must overcome. A similar descriptive approach has also been applied to association reactions (Wales 1987(Wales , 1996Ball et al. 1996;Wolynes 1996;Kumar et al. 2000). In these representations, a higher-dimensional problem must be compressed into a Abbreviations: N, number of subunits in a complete capsid; n, number of subunits in an intermediate; stat, the statistical factor reflecting assembly degeneracy over a whole capsid; DG contact , pairwise association energy between subunits; DG n,j , the overall association energy for the j-th intermediate of n subunits; P, probability of the specified intermediate; m, a weighting factor for reaction chemistry; s, the statistical factor reflecting assembly degeneracy for a specific reaction; f, the forward reaction rate for a specified reaction, a function of s and a microscopic rate; b, the backward rate for a specified reaction, a function of f and DG n,j ; t, time.Article and publication are at

We related events in immunohistochemical studies on formalin-fixed, paraffin-embedded preparations of 146 colorectal cancer patients. One hundred (68.5%) revealed cytoplasmic staining and only 3 of the 100 showed membranous staining. Western blot analysis revealed proteins with molecular masses of 185 and 155 kD following immunohistochemical membranous and cytoplasmic staining, but only a 155-kD protein following cytoplasmic staining. Amplification of the c-erbB-2 gene was detected in 2 of 44 cases (4.5%). Cytoplasmic c-erbB-2 overexpression correlated with tumor size (p < 0.01), subserosal invasion (p < 0.05), liver metastasis (p < 0.01) and Dukes’ classification. Overall survival rates and survival rates for Dukes’B patients were significantly lower in the group with cytoplasmic c-erbB-2 overexpression than in the group without cytoplasmic c-erbB-2 overexpression. Multivariate regression analysis showed cytoplasmic c-erbB-2 overexpression to be an independent prognostic factor for colorectal cancer. These results suggest that overexpression of cytoplasmic c-erbB-2 protein plays an important role in the progression of colorectal cancer and is considered to be an independent prognostic indicator of this lesion.

Matrix metalloproteinases (MMP), such as 72 kDa type IV collagenase (MMP-2) and 92 kDa type IV collagenase (MMP-9), play an important role in tumor invasion and metastasis. Tissue inhibitors of metalloproteinases (TIMP-1 and TIMP-2) are specific inhibitors of MMP. To evaluate the expression of TIMP-1, TIMP-2, MMP-2, and MMP-9 in human colorectal cancer, surgical specimens of primary colorectal cancer (66 cases) and liver metastases (10 cases) were examined by Northern and dot-blot hybridization. The levels of TIMP-1, MMP-2 and MMP-9 mRNA were significantly higher in primary colorectal cancers than in their adjacent normal tissues, and those of the mRNAs for all four genes were significantly higher in liver metastases than in normal colorectal tissues. Higher levels of TIMP-1 mRNA were positively correlated with lymph node metastasis and the five-year survival, and higher levels of TIMP-1 and TIMP-2 mRNA were positively correlated with the Dukes classification. Our findings suggest that the expression of TIMP-1 and TIMP-2 is closely correlated with the progression of human colorectal cancer.

We have previously shown that GAA trinucleotide repeats have undergone significant expansion in the human genome. Here we present the analysis of the length distribution of all 10 nonredundant trinucleotide repeat motifs in 20 complete eukaryotic genomes (6 mammalian, 2 nonmammalian vertebrates, 4 arthropods, 4 fungi, and 1 each of nematode, amoebozoa, alveolate, and plant), which showed that the abundance of large expansions of GAA trinucleotide repeats is specific to mammals. Analysis of human-chimpanzee-gorilla orthologs revealed that loci with large expansions are species-specific and have occurred after divergence from the common ancestor. PCR analysis of human controls revealed large expansions at multiple human (GAA)(30+) loci; nine loci showed expanded alleles containing >65 triplets, analogous to disease-causing expansions in Friedreich ataxia, including two that are in introns of genes of unknown function. The abundance of long GAA trinucleotide repeat tracts in mammalian genomes represents a significant mutation potential and source of interindividual variability.