Cell differentiation is remarkably stable but can be reversed by somatic cell nuclear transfer, cell fusion, and iPS. Nuclear transfer to amphibian oocytes provides a special opportunity to test ...transcriptional reprogramming without cell division. We show here that, after nuclear transfer to amphibian oocytes, mitotic chromatin is reprogrammed up to 100 times faster than interphase nuclei. We find that, as cells traverse mitosis, their genes pass through a temporary phase of unusually high responsiveness to oocyte reprogramming factors (mitotic advantage). Mitotic advantage is not explained by nuclear penetration, DNA modifications, histone acetylation, phosphorylation, methylation, nor by salt soluble chromosomal proteins. Our results suggest that histone H2A deubiquitination may account, at least in part, for the acquisition of mitotic advantage. They support the general principle that a temporary access of cytoplasmic factors to genes during mitosis may facilitate somatic cell nuclear reprogramming and the acquisition of new cell fates in normal development.
Circulating tumor cells (CTCs) are shed into the bloodstream from primary and metastatic tumor deposits. Their isolation and analysis hold great promise for the early detection of invasive cancer and ...the management of advanced disease, but technological hurdles have limited their broad clinical utility. We describe an inertial focusing-enhanced microfluidic CTC capture platform, termed "CTC-iChip," that is capable of sorting rare CTCs from whole blood at 10(7) cells/s. Most importantly, the iChip is capable of isolating CTCs using strategies that are either dependent or independent of tumor membrane epitopes, and thus applicable to virtually all cancers. We specifically demonstrate the use of the iChip in an expanded set of both epithelial and nonepithelial cancers including lung, prostate, pancreas, breast, and melanoma. The sorting of CTCs as unfixed cells in solution allows for the application of high-quality clinically standardized morphological and immunohistochemical analyses, as well as RNA-based single-cell molecular characterization. The combination of an unbiased, broadly applicable, high-throughput, and automatable rare cell sorting technology with generally accepted molecular assays and cytology standards will enable the integration of CTC-based diagnostics into the clinical management of cancer.
Nuclear transfer to oocytes is an efficient way to transcriptionally reprogram somatic nuclei, but its mechanisms remain unclear. Here, we identify a sequence of molecular events that leads to rapid ...transcriptional reprogramming of somatic nuclei after transplantation to Xenopus oocytes. RNA-seq analyses reveal that reprogramming by oocytes results in a selective switch in transcription toward an oocyte rather than pluripotent type, without requiring new protein synthesis. Time-course analyses at the single-nucleus level show that transcriptional reprogramming is induced in most transplanted nuclei in a highly hierarchical manner. We demonstrate that an extensive exchange of somatic- for oocyte-specific factors mediates reprogramming and leads to robust oocyte RNA polymerase II binding and phosphorylation on transplanted chromatin. Moreover, genome-wide binding of oocyte-specific linker histone B4 supports its role in transcriptional reprogramming. Thus, our study reveals the rapid, abundant, and stepwise loading of oocyte-specific factors onto somatic chromatin as important determinants for successful reprogramming.
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•Xenopus oocytes induce an oocyte transcription pattern in mouse nuclei in 2 days•Reprogramming requires a switch from somatic to oocyte transcriptional components•Unusually high amounts of oocyte-derived RNA polymerase II drive reprogramming•The pattern of oocyte linker histone binding to somatic chromatin is revealed
Jullien et al. identify a sequence of events leading to rapid transcriptional reprogramming of somatic nuclei after their transplantation to Xenopus oocytes. Transcription is switched selectively to an oocyte-type program in a process involving stepwise loading of oocyte-specific factors onto transplanted chromatin.
We introduce AutoEnRichness, a hybrid approach that combines empirical and analytical strategies to determine the richness of galaxy clusters (in the redshift range of \(0.1 \leq z \leq 0.35\)) using ...photometry data from the Sloan Digital Sky Survey Data Release 16, where cluster richness can be used as a proxy for cluster mass. In order to reliably estimate cluster richness, it is vital that the background subtraction is as accurate as possible when distinguishing cluster and field galaxies to mitigate severe contamination. AutoEnRichness is comprised of a multi-stage machine learning algorithm that performs background subtraction of interloping field galaxies along the cluster line-of-sight and a conventional luminosity distribution fitting approach that estimates cluster richness based only on the number of galaxies within a magnitude range and search area. In this proof-of-concept study, we obtain a balanced accuracy of \(83.20\) per cent when distinguishing between cluster and field galaxies as well as a median absolute percentage error of \(33.50\) per cent between our estimated cluster richnesses and known cluster richnesses within \(r_{200}\). In the future, we aim for AutoEnRichness to be applied on upcoming large-scale optical surveys, such as the Legacy Survey of Space and Time and \(\textit{Euclid}\), to estimate the richness of a large sample of galaxy groups and clusters from across the halo mass function. This would advance our overall understanding of galaxy evolution within overdense environments as well as enable cosmological parameters to be further constrained.
The T790M gatekeeper mutation in the EGFR is acquired by some EGFR-mutant non-small cell lung cancers (NSCLC) as they become resistant to selective tyrosine kinase inhibitors (TKI). As ...third-generation EGFR TKIs that overcome T790M-associated resistance become available, noninvasive approaches to T790M detection will become critical to guide management.
As part of a multi-institutional Stand-Up-To-Cancer collaboration, we performed an exploratory analysis of 40 patients with EGFR-mutant tumors progressing on EGFR TKI therapy. We compared the T790M genotype from tumor biopsies with analysis of simultaneously collected circulating tumor cells (CTC) and circulating tumor DNA (ctDNA).
T790M genotypes were successfully obtained in 30 (75%) tumor biopsies, 28 (70%) CTC samples, and 32 (80%) ctDNA samples. The resistance-associated mutation was detected in 47% to 50% of patients using each of the genotyping assays, with concordance among them ranging from 57% to 74%. Although CTC- and ctDNA-based genotyping were each unsuccessful in 20% to 30% of cases, the two assays together enabled genotyping in all patients with an available blood sample, and they identified the T790M mutation in 14 (35%) patients in whom the concurrent biopsy was negative or indeterminate.
Discordant genotypes between tumor biopsy and blood-based analyses may result from technological differences, as well as sampling different tumor cell populations. The use of complementary approaches may provide the most complete assessment of each patient's cancer, which should be validated in predicting response to T790M-targeted inhibitors.
Recent discoveries suggest that the candidate superphyla Patescibacteria and DPANN constitute a large fraction of the phylogenetic diversity of Bacteria and Archaea. Their small genomes and limited ...coding potential have been hypothesized to be ancestral adaptations to obligate symbiotic lifestyles. To test this hypothesis, we performed cell-cell association, genomic, and phylogenetic analyses on 4,829 individual cells of Bacteria and Archaea from 46 globally distributed surface and subsurface field samples. This confirmed the ubiquity and abundance of Patescibacteria and DPANN in subsurface environments, the small size of their genomes and cells, and the divergence of their gene content from other Bacteria and Archaea. Our analyses suggest that most Patescibacteria and DPANN in the studied subsurface environments do not form specific physical associations with other microorganisms. These data also suggest that their unusual genomic features and prevalent auxotrophies may be a result of ancestral, minimal cellular energy transduction mechanisms that lack respiration, thus relying solely on fermentation for energy conservation.
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