Passage Overexpression of the miR-17~92 gene cluster occurs in many human cancers involving c-Myc, a gene coding for a transcription factor that regulates expression of proteins such as p21, an inhibitor of cell cycle progression. miR-17~92 encodes six distinct microRNAs (miRNAs) that can be divided into four families based on sequence identity at positions 2-7 (Table 1).Table 1 Sequence Identities of miRNA Families Encoded by the mir-17∼92 Gene Cluster Experiment 1Two independent, diffuse B-cell lymphoma cell lines were used to study the functional relationship between miR-17~92 transcripts and c-Myc in cancer pathogenesis. The first cell line was derived from Eμ-Myc mice (Eμ), which overexpress a c-Myc transgene under the control of a B-cell-specific enhancer that induces early development of B-cell lymphoma. The second cell line was derived from Eμ-Myc mice carrying a miR-17∼92 knockout allele (Δ/Δ). As shown in Figure 1, researchers measured the growth of both cell lines, as well as that of Δ/Δ cells infected with a retrovirus expressing the entire miR-17∼92 cluster (Δ/Δ + miR-17∼92). Figure 1 Growth curves of Eμ cells, Δ/Δ cells, and Δ/Δ + miR-17∼92 cells (error bars = standard deviation)Experiment 2To assess the relative contribution of each miRNA to the oncogenic ability of the miR-17∼92 cluster, Δ/Δ cell lines were transduced with an empty vector or with miR-17∼92 mutant allele constructs that were engineered to lack expression of miRNA from at least one of the four families. In each of the constructs, the family that was knocked out of the vector is indicated by the Δ symbol. After verifying that the transduced cells correctly expressed the desired miRNA, the fraction of apoptotic cells in each line was measured (Figure 2). Figure 2 Percentage of apoptotic cells in Eμ cells and Δ/Δ cells transduced with the indicated miR-17~92 construct (error bars = standard deviation) Adapted from Mu P, Han YC, Betel D, et al. Genetic dissection of the miR-17~92 cluster of microRNAs in Myc-induced B-cell lymphomas. Genes Dev. 2009;23(24):2806-11. -Based on Figure 2, which family from the miR-17~92 gene cluster is expected to contribute the most to cancer progression?

Question

Quizplus · Accepted Answer

11ecba2d_0f62_f268_a3bf_fbabf31956b5_MD0008_00 Apoptosis, or the programmed death of preexisting cells, limits the rate of cell proliferation. However, cancerous tumors develop as cells grow/divide uncontrollably and fail to follow predetermined apoptotic pathways. As a result, the fraction of apoptotic cells within a group of cancer cells is markedly decreased (inhibited). In Experiment 2, the researchers wanted to identify which miRNA family from the miR-17~92 cluster contributed the most to apoptosis inhibition (ie, contributed most to cancer progression). First, a series of miR-17∼92 mutant allele constructs engineered to lack expression of specific miRNA families (indicated by Δ) were transduced into Eμ-Myc cells with the entire miR-17~92 cluster knocked out (Δ/Δ). The fraction of apoptotic cells was then measured in these cell lines. As shown in Figure 2, out of all the transduced cell lines, the cell line expressing all miR-17~92 families except miR-19 (Δ/Δ + 17~92 (Δ19)) had the highest percentage of apoptotic cells. This result signifies that removal of only miR-19 from the miR-17~92 cluster promotes apoptosis. Accordingly, the miR-19 family is therefore the most essential to cancer progression as its expression suppresses apoptosis. (Choice A) To precisely conclude whether miR-17 has or lacks oncogenic properties, the percentage of apoptotic cells would have to be measured in Eμ-Myc cell lines lacking miR-17 only (Δ/Δ + Δ17), not in cells lacking both miR-17 and miR-18 (shown in Figure 2). (Choices B and D) The cell line that lacks only miR-18 and the cell line that lacks only miR-92 still show a low percentage of apoptotic cells (characteristic of cancer progression). This finding means that both miR-18 and miR-92 do not contribute as much as miR-19 to promoting decreased apoptosis in cancer progression. Educational objective: Apoptosis is the programmed and controlled death of aged, unnecessary, or damaged cells. Cancerous cells exhibit rapid and uncontrolled proliferation, which is caused by cell growth that outpaces normal apoptosis or by dysfunction of the apoptotic process itself.

Passage Overexpression of the MiR-17~92 Gene Cluster Occurs in Many Human