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The let-7 microRNA Family Regulates RAS: Implications for Development and Oncogenesis

cancer samples.

To test this hypothesis, both RNA and protein were isolated from three pairs of lung squamous cell carcinoma and NAT using the mirVana PARIS Kit, so that RAS protein and RNA levels and let-7 miRNA levels could be measured in the same samples. In all cases, RAS protein levels in tumor cells were at least ten times higher than in the corresponding NAT (Figure 3C, top), and let-7 miRNA levels in the tumor were 4- to 8-fold lower than in the corresponding NAT (Figure 3C, middle). This result indicates that RAS protein expression correlates better with miRNA expression levels than with RAS mRNA in lung.

Figure 3. let-7 is Poorly Expressed in Human Lung Tumors. (A) The mirVana miRNA Isolation Kit (Ambion) was used to isolate miRNA from 21 breast, colon, and lung tumors, as well as from associated normal adjacent tissue (NAT). Microarray analysis using 167 miRNA probes (mirVana miRNA Probe Set, Ambion) was used to examine let-7 miRNA expression profiles (tumor relative to NAT) in these samples. After labeling tumor miRNA samples with Cy3 (Amersham) and NAT miRNA samples with Cy5 (mirVana miRNA Labeling Kit, Ambion), the microarray was hybridized for 14 h. Scanning was performed with the GenePix 4000B (Axon) (B) Northern analysis was performed using paired RNA preparations from two of the patients from Figure 3A and a radiolabeled probe specific for let-7c and 5S rRNA (control). (C) The mirVana PARIS Kit (Ambion) was used to isolate protein and RNA fr
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