Enhancement of cell proliferation in various mammalian cell lines by gene insertion of a cyclin-dependent kinase homolog

<p>Abstract</p> <p>Background</p> <p>Genomics tools, particularly DNA microarrays, have found application in a number of areas including gene discovery and disease characterization. Despite the vast utility of these tools, little work has been done to explore the basis of distinct cellular properties, especially those important to biotechnology such as growth. And so, with the intent of engineering cell lines by manipulating the expression of these genes, anchorage-independent and anchorage-dependent HeLa cells, displaying markedly different growth characteristics, were analyzed using DNA microarrays.</p> <p>Results</p> <p>Two genes, cyclin-dependent kinase like 3 (<it>cdkl3</it>) and cytochrome c oxidase subunit (<it>cox15</it>), were up-regulated in the faster growing, anchorage-independent (suspension) HeLa cells relative to the slower growing, anchorage-dependent (attached) HeLa cells. Enhanced expression of either gene in the attached HeLa cells resulted in elevated cell proliferation, though insertion of <it>cdkl3 </it>had a greater impact than that of <it>cox15</it>. Moreover, flow cytometric analysis indicated that cells with an insert of <it>cdkl3 </it>were able to transition from the G0/G1 phases to the S phase faster than control cells. In turn, expression of <it>cox15 </it>was seen to increase the maximum viable cell numbers achieved relative to the control, and to a greater extent than <it>cdkl3</it>. Quantitatively similar results were obtained with two Human Embryonic Kidney-293 (HEK-293) cell lines and a Chinese Hamster Ovary (CHO) cell line. Additionally, HEK-293 cells secreting adipocyte complement-related protein of 30 kDa (acrp30) exhibited a slight increase in specific protein production and higher total protein production in response to the insertion of either <it>cdkl3 </it>or <it>cox15</it>.</p> <p>Conclusion</p> <p>These results are consistent with previous studies on the functionalities of <it>cdkl3 </it>and <it>cox15</it>. For instance, the effect of <it>cdkl3 </it>on cell growth is consistent with its homology to the <it>cdk3 </it>gene which is involved in G1 to S phase transition. Likewise, the increase in cell viability due to <it>cox15 </it>expression is consistent with its role in oxidative phosphorylation as an assembly factor for cytochrome c oxidase and its involvement removing apoptosis-inducing oxygen radicals. Collectively, the present study illustrates the potential of using microarray technology to identify genes influential to specific cellular processes with the possibility of engineering cell lines as desired to meet production needs.</p

A perspective on microarrays: current applications, pitfalls, and potential uses

Abstract With advances in robotics, computational capabilities, and the fabrication of high quality glass slides coinciding with increased genomic information being available on public databases, microarray technology is increasingly being used in laboratories around the world. In fact, fields as varied as: toxicology, evolutionary biology, drug development and production, disease characterization, diagnostics development, cellular physiology and stress responses, and forensics have benefiting from its use. However, for many researchers not familiar with microarrays, current articles and reviews often address neither the fundamental principles behind the technology nor the proper designing of experiments. Although, microarray technology is relatively simple, conceptually, its practice does require careful planning and detailed understanding of the limitations inherently present. Without these considerations, it can be exceedingly difficult to ascertain valuable information from microarray data. Therefore, this text aims to outline key features in microarray technology, paying particular attention to current applications as outlined in recent publications, experimental design, statistical methods, and potential uses. Furthermore, this review is not meant to be comprehensive, but rather substantive; highlighting important concepts and detailing steps necessary to conduct and interpret microarray experiments. Collectively, the information included in this text will highlight the versatility of microarray technology and provide a glimpse of what the future may hold.</p