Many biological processes are controlled by intricate networks of transcriptional regulators. With the development of microarray technology, transcriptional changes can be examined at the whole-genome level. However, such analysis often lacks information on the hierarchical relationship between components of a given system. Systemic acquired resistance (SAR) is an inducible plant defense response involving a cascade of transcriptional events induced by salicylic acid through the transcription cofactor NPR1. To identify additional regulatory nodes in the SAR network, we performed microarray analysis on Arabidopsis plants expressing the NPR1-GR (glucocorticoid receptor) fusion protein.Since nuclear translocation of NPR1-GR requires dexamethasone, we were able to control NPR1-dependent transcription and identify direct transcriptional targets of NPR1. We show that NPR1 directly upregulates the expression of eight WRKY transcription factor genes. This large family of 74 transcription factors has been implicated in various defense responses, but no specific WRKY factor has been placed in the SAR network. Identification of NPR1-regulated WRKY factors allowed us to perform in-depth genetic analysis on a small number of WRKY factors and test well-defined phenotypes of single and double mutants associated with NPR1. Among these WRKY factors we found both positive and negative regulators of SAR. This genomics-directed approach unambiguously positioned five WRKY factors in the complex transcriptional regulatory network of SAR. Our work not only discovered new transcription regulatory components in the signaling network of SAR but also demonstrated that functional studies of large gene families have to take into consideration sequence similarity as well as the expression patterns of the candidates.
Purpose: To discover diagnostic biomarkers associated with early-stage non-small cell lung cancer (NSCLC), we searched for autoantibodies preferentially present in stage I patients compared with patients with advanced-stage disease. Here we describe an autoantibody against complement factor H (CFH) and this autoantibody's association with early-stage NSCLC.Experimental Design: Immunoblots were used to detect autoantibodies in the sera of stage I NSCLC patients. An autoantibody recognizing a 150 kDa protein was discovered, and the protein was identified by mass spectrometry. The association of the autoantibody with early-stage disease was suggested by the results of immunoblot analysis with sera from 28 stage I patients and 28 stage III/IV patients. This association was confirmed by protein microarray of sera from 125 NSCLC patients of all stages as well as 125 controls matched by age, gender, and smoking history.Results: The immunoreactive protein was identified as CFH. By immunoblot analysis, anti-CFH autoantibody was found in 50% of stage I NSCLC patients and 11% of late-stage NSCLC patients (P = 0.003). By protein microarray analysis, patients with stage I NSCLC had a significantly higher incidence of anti-CFH antibody than those with late-stage NSCLC (P = 0.0051). The percentage of sera with a positive level of CFH autoantibody was 30.4% in stage I, 21.1% in stage II, 12.5% in stage III, 7.4% in stage IV, and 8.0% in the control group.Conclusions: These findings suggest that in patients with NSCLC, CFH autoantibody is a molecular marker associated with early-stage disease. Clin Cancer Res; 16(12); 3226-31. ©2010 AACR.Patients with non-small cell lung cancer (NSCLC) typically present with advanced-stage disease, and almost all will eventually die with metastasis. Even patients classified as having stage I disease have a 5-year survival rate of <60%, suggesting undetected micrometastasis at the time of presentation (1, 2). Unfortunately, attempts to improve outcome through early detection efforts using chest radiographs, sputum cytology, and more recently, low-dose computed tomography (CT) have failed to show a reduction in mortality. New strategies that make use of our continually improving understanding of oncogenesis and the process of metastasis are needed if early detection efforts are to be successful.One approach to early detection is the discovery of novel biomarkers that could not only identify patients with lung cancer, but could assist in appropriate staging. Autoantibodies are one class of potentially useful serum biomarkers as there is a low incidence of cancer-associated autoantibodies in healthy individuals (3). In addition, the autoantibody response elicited against a tumor-associated antigen may elucidate a unique therapeutic target, albeit in a select group of cancer patients. Here we report the discovery of an autoantibody to complement factor H (CFH), one of the complement inhibitory proteins on tumor cells, and show its association with early-stage NSCLC. Materials and Methods Pati...
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