The prevailing view that the evolution of cells in a tumor is driven by Darwinian selection has never been rigorously tested. Because selection greatly affects the level of intratumor genetic diversity, it is important to assess whether intratumor evolution follows the Darwinian or the non-Darwinian mode of evolution. To provide the statistical power, many regions in a single tumor need to be sampled and analyzed much more extensively than has been attempted in previous intratumor studies. Here, from a hepatocellular carcinoma (HCC) tumor, we evaluated multiregional samples from the tumor, using either whole-exome sequencing (WES) (n = 23 samples) or genotyping (n = 286) under both the infinite-site and infinite-allele models of population genetics. In addition to the many single-nucleotide variations (SNVs) present in all samples, there were 35 “polymorphic” SNVs among samples. High genetic diversity was evident as the 23 WES samples defined 20 unique cell clones. With all 286 samples genotyped, clonal diversity agreed well with the non-Darwinian model with no evidence of positive Darwinian selection. Under the non-Darwinian model, MALL (the number of coding region mutations in the entire tumor) was estimated to be greater than 100 million in this tumor. DNA sequences reveal local diversities in small patches of cells and validate the estimation. In contrast, the genetic diversity under a Darwinian model would generally be orders of magnitude smaller. Because the level of genetic diversity will have implications on therapeutic resistance, non-Darwinian evolution should be heeded in cancer treatments even for microscopic tumors.

In actinomycetes, the onset of secondary metabolite biosynthesis is often triggered by the quorum-sensing signal ␥-butyrolactones (GBLs) via specific binding to their cognate receptors. However, the presence of multiple putative GBL receptor homologues in the genome suggests the existence of an alternative regulatory mechanism. Here, in the model streptomycete Streptomyces coelicolor, ScbR2 (SCO6286, a homologue of GBL receptor) is shown not to bind the endogenous GBL molecule SCB1, hence designated "pseudo" GBL receptor. Intriguingly, it could bind the endogenous antibiotics actinorhodin and undecylprodigiosin as ligands, leading to the derepression of KasO, an activator of a cryptic type I polyketide synthase gene cluster. Likewise, JadR2 is also a putative GBL receptor homologue in Streptomyces venezuelae, the producer of chloramphenicol and cryptic antibiotic jadomycin. It is shown to coordinate their biosynthesis via direct repression of JadR1, which activates jadomycin biosynthesis while repressing chloramphenicol biosynthesis directly. Like ScbR2, JadR2 could also bind these two disparate antibiotics, and the interactions lead to the derepression of jadR1. The antibiotic responding activities of these pseudo GBL receptors were further demonstrated in vivo using the lux reporter system. Overall, these results suggest that pseudo GBL receptors play a novel role to coordinate antibiotic biosynthesis by binding and responding to antibiotics signals. Such an antibiotic-mediated regulatory mechanism could be a general strategy to coordinate antibiotic biosynthesis in the producing bacteria.

In bacteria, many ''atypical'' response regulators (ARRs) lack the conserved residues important for phosphorylation by which typical response regulators switch their output response, suggesting the existence of alternative regulatory mechanisms. However, such mechanisms have not been established. JadR1, an OmpR-type ARR of Streptomyces venezuelae, appears to activate the transcription of jadomycin B (JdB) biosynthetic genes while repressing its own gene. JadR1 activities were inhibited in cells induced to produce JdB, which was found to bind directly to the N-terminal receiver domain of JadR1, causing JadR1 to dissociate from target promoters. The activity of a NarL-type ARR, RedZ, that regulates production of another antibiotic was likewise modulated by the end product (undecylprodigisines), implying that end-product-mediated control of antibiotic pathway-specific ARRs may be widespread. These results could prove relevant to knowledge-based improvements in yield of commercially important antibiotics.JadR1 ͉ ligand

Microbes live mostly in a social community rather than in a planktonic state. Such communities have complex spatiotemporal patterns that require intercellular communication to coordinate gene expression. Here, we demonstrate that Cryptococcus neoformans, a model eukaryotic pathogen, responds to an extracellular signal in constructing its colony morphology. The signal that directs this community behavior is not a molecule of low molecular weight like pheromones or quorum-sensing molecules but a secreted protein. Znf2, a master regulator of morphogenesis in Cryptococcus, is necessary and sufficient for the production of this signal protein. Cfl1, a prominent Znf2-downstream adhesion protein (adhesin), was identified to be responsible for the paracrine communication. Consistent with its role in communication, Cfl1 is highly induced during mating colony differentiation, and some of the Cfl1 proteins undergo shedding and are released from the cell wall. The released Cfl1 is enriched in the extracellular matrix and acts as an autoinduction signal to stimulate neighboring cells to phenocopy Cfl1-expressing cells via the filamentationsignaling pathway. We further demonstrate the importance of an unannotated and yet conserved domain in Cfl1's signaling activity. Although adhesion proteins have long been considered to be mediators of microbial pathogenicity and the structural components of biofilms, our work presented here provides the direct evidence supporting the signaling activation by microbial adhesion/matrix proteins.fungal community behavior | extracellular matrix signaling | fungal dimorphism | flocculin

Esophageal carcinoma is characterized by a widely ranged incidence variation among the different geographic regions. Anyang is a county in Henan Province of North China with the highest prevalence of esophageal carcinoma. Human papillomavirus (HPV) infection has been linked to the etiology of esophageal cancer in this area. In this study, we investigated correlations of the polymorphisms at low molecular weight polypeptide (LMP) and transporters with antigen processing (TAP) genes, with the risk of esophageal carcinoma. DNA extracted from either tumor specimens or esophageal epithelial cells was used to test HPV infection. Peripheral blood lymphocyte DNA was used for LMP/TAP genotyping. Polymerase chain reaction was performed to analyze HPV infection and LMP/TAP gene polymorphisms. The combined effect of LMP/TAP gene polymorphisms and HPV infection on esophageal carcinoma was analyzed by using unconditional logistic regression models. The TAP2 codons 379 isoleucine carriers and LMP7 codons 145 lysine carriers were found to be more susceptible to esophageal carcinoma (OR = 2.74, 95% CI = 1.15-6.49, P = 0.023 for TAP2; OR = 2.19, 95% CI = 1.09-4.37, P = 0.027 for LMP7). Patients carrying homozygous LMP7/TAP2 haplotype C, which contained the glutamine at LMP7 codons 145 and the isoleucine at TAP2 codons 379, were more prone to develop esophageal carcinoma (OR = 2.96, 95% CI = 1.13-7.81, P = 0.027). An additive effect on the risk of esophageal carcinoma development was found among individuals carrying LMP7/TAP2 haplotype C and infected by HPV (OR = 4.33, 95% CI = 2.53-7.42, P < 0.0001). LMP7/TAP2 haplotype C may act as the risk factor in esophageal carcinoma development and it may influence the tumorigenesis in HPV infected individuals.

Cancer stem cells are implicated in tumor progression, metastasis, and recurrence, although the exact mechanisms remain poorly understood. Here, we show that the expression of cellular prion protein (PrPc, PRNP) is positively correlated with an increased risk of metastasis in colorectal cancer. PrPc defines a subpopulation of CD44-positive cancer stem cells that contributes to metastatic capacity.

In the human fungal pathogen Cryptococcus neoformans, sex can benefit its pathogenicity through production of meiospores, which are believed to offer both physical and meiosis-created lineage advantages for its infections. Cryptococcus sporulation occurs following two parallel events, meiosis and differentiation of the basidium, the characteristic sexual structure of the basidiomycetes. However, the circuit integrating these events to ensure subsequent sporulation is unclear. Here, we show the spatiotemporal coordination of meiosis and basidial maturation by visualizing event-specific molecules in developing basidia defined by a quantitative approach. Monitoring of gene induction timing together with genetic analysis reveals co-regulation of the coordinated events by a shared regulatory program. Two RRM family regulators, Csa1 and Csa2, are crucial components that bridge meiosis and basidial maturation, further determining sporulation. We propose that the regulatory coordination of meiosis and basidial development serves as a determinant underlying the production of infectious meiospores in C. neoformans.

Sexual reproduction in an environmental pathogen helps maximize its lineage fitness to changing environment and the host. For the fungal pathogen Cryptococcus neoformans, sexual reproduction is proposed to have yielded hyper virulent and drug resistant variants. The life cycle of this pathogen commences with mating, followed by the yeast-hypha transition and hyphal growth, and it concludes with fruiting body differentiation and sporulation. How these sequential differentiation events are orchestrated to ensure developmental continuality is enigmatic. Here we revealed the genetic network of the yeast-to-hypha transition in Cryptococcus by analyzing transcriptomes of populations with a homogeneous morphotype generated by an engineered strain. Among this network, we found that a Pumilio-family protein Pum1 and the matricellular signal Cfl1 represent two major parallel circuits directing the yeast-hypha transition. Interestingly, only Pum1 coordinates the sequential morphogenesis events during a-α bisexual and α unisexual reproduction. Pum1 initiates the yeast-to-hypha transition, partially through a novel filament-specific secretory protein Fas1; Pum1 is also required to sustain hyphal growth after the morphological switch. Furthermore, Pum1 directs subsequent differentiation of aerial hyphae into fruiting bodies in both laboratory and clinical isolates. Pum1 exerts its control on sexual reproduction partly through regulating the temporal expression of Dmc1, the meiosis-specific recombinase. Therefore, Pum1 serves a pivotal role in bridging post-mating morphological differentiation events with sexual reproduction in Cryptococcus. Our findings in Cryptococcus illustrate how an environmental pathogen can ensure the completion of its life cycle to safeguard its long-term lineage success.