scholarly journals An Integrated Preprocessing Approach for Exploring Single-Cell Gene Expression in Rare Cells

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Junyi Shang ◽  
David Welch ◽  
Manuela Buonanno ◽  
Brian Ponnaiya ◽  
Guy Garty ◽  
...  
Keyword(s):  
Gene Expression ◽  
Single Cell ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Bystander Effect ◽  
Kinase Inhibitor ◽  
Digital Pcr ◽  
Lung Fibroblasts ◽  
Gene Expressions ◽  
Rare Cells

AbstractExploring the variability in gene expressions of rare cells at the single-cell level is critical for understanding mechanisms of differentiation in tissue function and development as well as for disease diagnostics and cancer treatment. Such studies, however, have been hindered by major difficulties in tracking the identity of individual cells. We present an approach that combines single-cell picking, lysing, reverse transcription and digital polymerase chain reaction to enable the isolation, tracking and gene expression analysis of rare cells. The approach utilizes a photocleavage bead-based microfluidic device to synthesize and deliver stable cDNA for downstream gene expression analysis, thereby allowing chip-based integration of multiple reactions and facilitating the minimization of sample loss or contamination. The utility of the approach was demonstrated with QuantStudio digital PCR by analyzing the radiation and bystander effect on individual IMR90 human lung fibroblasts. Expression levels of the Cyclin-dependent kinase inhibitor 1a (CDKN1A), Growth/differentiation factor 15 (GDF15), and Prostaglandin-endoperoxide synthase 2 (PTGS2) genes, previously shown to have different responses to direct and bystander irradiation, were measured across individual control, microbeam-irradiated or bystander IMR90 cells. In addition to the confirmation of accurate tracking of cell treatments through the system and efficient analysis of single-cell responses, the results enable comparison of activation levels of different genes and provide insight into signaling pathways within individual cells.

scholarly journals Using single-cell transcriptomics to understand functional states and interactions in microbial eukaryotes

2019 ◽  
Vol 374 (1786) ◽  
pp. 20190098 ◽  
Author(s):  
Chuan Ku ◽  
Arnau Sebé-Pedrós
Keyword(s):  
Gene Expression ◽  
Single Cell ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Interspecific Interactions ◽  
Single Cells ◽  
Life Cycles ◽  
Modern Biology ◽  
Microbial Eukaryotes ◽  
Eukaryotic Microorganisms

Understanding the diversity and evolution of eukaryotic microorganisms remains one of the major challenges of modern biology. In recent years, we have advanced in the discovery and phylogenetic placement of new eukaryotic species and lineages, which in turn completely transformed our view on the eukaryotic tree of life. But we remain ignorant of the life cycles, physiology and cellular states of most of these microbial eukaryotes, as well as of their interactions with other organisms. Here, we discuss how high-throughput genome-wide gene expression analysis of eukaryotic single cells can shed light on protist biology. First, we review different single-cell transcriptomics methodologies with particular focus on microbial eukaryote applications. Then, we discuss single-cell gene expression analysis of protists in culture and what can be learnt from these approaches. Finally, we envision the application of single-cell transcriptomics to protist communities to interrogate not only community components, but also the gene expression signatures of distinct cellular and physiological states, as well as the transcriptional dynamics of interspecific interactions. Overall, we argue that single-cell transcriptomics can significantly contribute to our understanding of the biology of microbial eukaryotes. This article is part of a discussion meeting issue ‘Single cell ecology’.

scholarly journals Resolution of Cell Fate Decisions Revealed by Single-Cell Gene Expression Analysis from Zygote to Blastocyst

2010 ◽  
Vol 18 (4) ◽  
pp. 675-685 ◽  
Author(s):  
Guoji Guo ◽  
Mikael Huss ◽  
Guo Qing Tong ◽  
Chaoyang Wang ◽  
Li Li Sun ◽  
...  
Keyword(s):  
Gene Expression ◽  
Single Cell ◽  
Cell Fate ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Cell Fate Decisions ◽  
Cell Gene Expression ◽  
Cell Gene

scholarly journals Bovine lineage specification revealed by single-cell gene expression analysis from zygote to blastocyst†

2017 ◽  
Vol 97 (1) ◽  
pp. 5-17 ◽  
Author(s):  
Qingqing Wei ◽  
Liang Zhong ◽  
Shaopeng Zhang ◽  
Haiyuan Mu ◽  
Jinzhu Xiang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Single Cell ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Lineage Specification ◽  
Cell Gene Expression ◽  
Cell Gene

Bortezomib Is Effective for Down Regulation of Gene Transcriptions Specifically Expressed in Clonogenic Myeloma Side Population Cells

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3939-3939
Author(s):  
Miho Nara ◽  
Hiroyuki Tagawa ◽  
Kazuaki Teshima ◽  
Atsushi Watanabe ◽  
Mitsugu Ito ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cell ◽  
Cell Lines ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Kinase Inhibitor ◽  
Side Population ◽  
Aurora Kinase ◽  
Aurora Kinase B ◽  
Transporter Family

Abstract Abstract 3939 Background: Multiple myeloma (MM) is characterized by the accumulation of a population of malignant plasma cells within the bone marrow. Cytotoxic chemotherapy-based treatment is not curative, and the disease eventually recurs. Although currently available anti-MM strategies are effective at targeting the bulk of tumor cells, it is not clear that these agents are targeting the tumor-initiating subpopulation, or cancer stem cells. Side Population (SP) cells are an enriched source of cancer-initiating cells with stem cell properties, which have been identified in solid tumors, as well as in hematopoietic malignancies. SP cells express high levels of various members of the ABC transporter family, which are responsible for their drug resistance. A recent our work demonstrated that SP cells in MM have shown to exhibit stem cell like characteristics as well as high tumorigenicity. Therefore, it is worthy to identify gene/proteins specifically expressed in MM SP cells, which could be essential therapeutic targets. Purpose: The aim of this study was to identify genes and transcripts that could serve as molecular markers for targeting the MM SP cells, and to identify candidate agents for the MM SP cells. Experimental design: We used Hoechst 33342 dye to detect the MM SP in five MM cell lines (RPMI 8226, AMO1, KMS-12BM, KMS-11 and JJN3) and eight primary samples. We then tested whether the MM SP cells have stem-like characteristics and performed gene expression analysis to detect genes specifically expressed in the MM SP. On that basis, we tested candidate agents such as an aurora kinase inhibitor (VX-680), a histone methyltransferase inhibitor (DZNep), lenalidomide, thalidomide and a proteasome inhibitor (bortezomib) for their ability to target MM SP cells. Results: We found that clonogenic MM SP cells exhibit “stem cell-like” properties, including self renewal, differentiation and repopulation. Gene expression analysis of MM cell lines and primary samples revealed that, in SP cells, expression of genes related to G2/M phase (e.g. CDC2, CCNB1)-, microtubule attachment (e.g. BIRC5, CENPE, SKA1)-, mitosis or centrosomes (e.g. AURKB, KIF2C, KIF11, KIF15)-, proliferation (e.g. TOP2A, ASPM)-, polycomb (e.g. EZH2, EPC1)- and proteasomes(e.g. UBE2D3, UBE3C, PSMA5)- was significantly stronger in SP than non-SP cells. On that basis, we used VX-680, DZNep, lenalidomide, thalidomide and bortezomib against MM cells. Of these, bortezomib reduced the SP fraction most effectively due to its ability to reduce levels of target gene transcripts including phospho-histone H3, aurora kinase B and EZH2. Finally we tried to examine effects of those candidate agents to “clonogenic ability of SP”, and found that bortezomib possessed the most powerful effects for reduction of SP colonies. These results suggest that bortezomib has a broader range of targets than other agents and could include cell cycle, centrosome, polycomb and proteasome genes/proteins. Conclusion: Our findings are i) the first to identify genes specifically expressed in the MM SP, ii) the first to provide a rationale for treating MM using agents against genes and encoded proteins that are specifically expressed in MM SP cells. Disclosures: Iida: Janssen Pharmaceutical K.K.: Honoraria.

Sign in / Sign up

Export Citation Format

Share Document