Complementary DNA

About: Complementary DNA is a research topic. Over the lifetime, 55301 publications have been published within this topic receiving 2752650 citations. The topic is also known as: cDNA & DNA, Complementary.

SARPs: a family of secreted apoptosis-related proteins.

Abstract: Quiescent mouse embryonic C3H/10T½ cells are more resistant to different proapoptotic stimuli than are these cells in the exponential phase of growth. However, the exponentially growing 10T½ cells are resistant to inhibitors of RNA or protein synthesis, whereas quiescent cells die upon these treatments. Conditioned medium from quiescent 10T½ cells possesses anti-apoptotic activity, suggesting the presence of protein(s) that function as an inhibitor of the apoptotic program. Using differential display technique, we identified and cloned a cDNA designated sarp1 (secreted apoptosis-related protein) that is expressed in quiescent but not in exponentially growing 10T½ cells. Hybridization studies with sarp1 revealed two additional family members. Cloning and sequencing of sarp2 and sarp3 revealed 38% and 40% sequence identity to sarp1, respectively. Human breast adenocarcinoma MCF7 cells stably transfected with sarp1 or infected with SARP1-expressing adenovirus became more resistant, whereas cells transfected with sarp2 displayed increased sensitivity to different proapoptotic stimuli. Expression of sarp family members is tissue specific. sarp mRNAs encode secreted proteins that possess a cysteine-rich domain (CRD) homologous to the CRD of frizzled proteins but lack putative membrane-spanning segments. Expression of SARPs modifies the intracellular levels of β-catenin, suggesting that SARPs interfere with the Wnt–frizzled proteins signaling pathway.

A thymus-specific member of the HMG protein family regulates the human T cell receptor C alpha enhancer.

Abstract: The human T cell-specific transcription factor TCF-1 alpha plays a key role in the tissue-specific activation of the T cell receptor (TCR) C alpha enhancer and binds to pyrimidine-rich elements (5'-PyCTTTG-3') present in a variety of other T cell-specific control regions Using amino acid sequence information derived from the DNA affinity-purified protein, we have now isolated cDNA clones encoding TCF-1 alpha The TCF-1 alpha cDNA contains a single 68-amino-acid domain that is homologous to a region conserved among high-mobility group (HMG) and nonhistone chromosomal proteins Expression of full-length and mutant cDNA clones in bacteria reveal that the single HMG motif, which is predicted to contain two extended alpha-helical segments, is sufficient to direct the sequence-specific binding of TCF-1 alpha to DNA Northern blot experiments demonstrate further that TCF-1 alpha mRNA is highly tissue specific, found primarily in the thymus or T cell lines The immature CEM T cell line expresses relatively low levels of TCF-1 alpha mRNA, which are increased upon activation of these cells by phorbol esters Interestingly, the cloned TCF-1 alpha protein is a potent transcriptional activator of the human TCR alpha enhancer in nonlymphoid cell lines, whereas the activity of the endogenous protein in T cell lines is strongly dependent on an additional T cell-specific protein that interacts with the core enhancer TCF-1 alpha is currently unique among the newly emerging family of DNA-binding regulatory proteins that share the HMG motif in that it is a highly tissue-specific RNA polymerase II transcription factor

Engineering the largest RNA virus genome as an infectious bacterial artificial chromosome

Abstract: The construction of cDNA clones encoding large-size RNA molecules of biological interest, like coronavirus genomes, which are among the largest mature RNA molecules known to biology, has been hampered by the instability of those cDNAs in bacteria. Herein, we show that the application of two strategies, cloning of the cDNAs into a bacterial artificial chromosome and nuclear expression of RNAs that are typically produced within the cytoplasm, is useful for the engineering of large RNA molecules. A cDNA encoding an infectious coronavirus RNA genome has been cloned as a bacterial artificial chromosome. The rescued coronavirus conserved all of the genetic markers introduced throughout the sequence and showed a standard mRNA pattern and the antigenic characteristics expected for the synthetic virus. The cDNA was transcribed within the nucleus, and the RNA translocated to the cytoplasm. Interestingly, the recovered virus had essentially the same sequence as the original one, and no splicing was observed. The cDNA was derived from an attenuated isolate that replicates exclusively in the respiratory tract of swine. During the engineering of the infectious cDNA, the spike gene of the virus was replaced by the spike gene of an enteric isolate. The synthetic virus replicated abundantly in the enteric tract and was fully virulent, demonstrating that the tropism and virulence of the recovered coronavirus can be modified. This demonstration opens up the possibility of employing this infectious cDNA as a vector for vaccine development in human, porcine, canine, and feline species susceptible to group 1 coronaviruses.