Box plot of miRNA levels in ECs, KSHV-infected ECs, KS, and PEL. Sample classes are shown on the horizontal axis and relative miRNA levels (dCT) on the vertical axis. dCT indicates the miRNA abundance relative to U6 (set to 0). Lower dCT levels correspond to higher mRNA levels on a 2log scale. The line represents the median; boxes, 25% and 75% percentiles; whiskers, 1.5 times the box size. Outliers are indicated by an open circle. (A-F) Based on technical triplicate measurements of each sample and of multiple independent samples for each class. The number of biologic replicates were: ECs, n = 2; EC-KSHV, n = 4; KS, n = 4; PEL, n = 12. Thus, the variation depicted by the box-whisker plot reflects technical as well as biologic variation within each sample class. The P values in each case were less than or equal to .001. (G) Change in miRNA levels in KSHV latently infected ECs. The horizontal axis shows mean relative levels of dCT over n = 6 replicates (3 for uninfected and 3 for KSHV-infected ECs). Lower dCT levels correspond to higher mRNA levels on a log2 scale. The vertical axis shows the P value of the pairwise t test. miRNAs that were significant after adjustment for multiple comparisons25 are labeled. (H) Analysis of residuals for the same dataset. Residuals quantify how far any one actual data point deviates from the fitted prediction based on regression of all data points. Because in array analyses the majority of genes/miRNAs do not change, large residuals identify outliers, that is, those miRNAs that are changed between 2 samples. Analysis of residuals is a more robust and statistically sound technique than calculating the fold change between 2 data points for any one miRNA. Shown on the vertical axis are the residuals and on the horizontal axis the fitted values as predicted by robust linear regression analysis9 of KSHV-infected versus uninfected ECs. Significantly altered genes with residuals greater than 10 units are labeled. Additional changed genes are shown in Table S2.
