AP1 is a transcription factor and an important regulator of gene expression and cell proliferation. Dysregulation of AP1 function may lead to cancer. The fos gene codes for AP1. Below is an experiment in which the role of a micro‐RNA (miR‐7b) in the regulation of fos gene expression was studied. miR‐7b shows partial sequence complementarity to the 3′‐untranslated region of fos mRNA. A DNA fragment coding for miR‐7b RNA and another fragment coding for a micro‐RNA unrelated to fos mRNA were cloned into vectors, and the recombinant plasmids (designated si‐miR‐7b, samples 3 and 4, and si‐miR‐neg, samples 5 and 6, respectively, in Fig. 1) were introduced into mouse fibroblast cells. Non-transfected cells were used as controls (samples 1 and 2). Samples 2, 4, and 6, were treated with a chemical PMA that induces transcription of fos gene; samples 1, 3, and 5 were left untreated. Two hours after PMA treatment protein extracts were prepared from the cultures and were then subjected to Western blot analysis using anti‐Fos (upper panel, Fig. 1 a) and anti-actin (lower panel) antibodies. RNA was isolated from other cell cultures after 1-hour PMA treatment, separated on a special gel by electrophoresis. A labeled fos cDNA probe was added to hybridize the RNA and imaged. Bands indicate the hybridization of probe to RNA. 1) In your own words explain the purpose of the Western Blot and labeled cDNA probe in this experiment? 2) Why do you think they isolated RNA 1- hour after treatment and protein 2 hours after treatment 3) The following statements are related to the information presented in the description of the experiment. On the basis of the information given, select: A : if the statement is supported by the information given; B : if the statement is contradicted by the information given; C : if the statement is neither supported nor contradicted by the information given. a) miR‐7b has a general inhibitory effect on transcription in fibroblasts b) miR‐7b has a general inhibitory effect on translation in fibroblasts c) miR‐7b specifically blocks the expression of the Fos protein.
Question 5.
AP1 is a transcription factor and an important regulator of gene expression and cell proliferation. Dysregulation of AP1 function may lead to cancer.
The fos gene codes for AP1.
Below is an experiment in which the role of a micro‐RNA (miR‐7b) in the regulation of fos gene expression was studied. miR‐7b shows partial sequence complementarity to the 3′‐untranslated region of fos mRNA.
A DNA fragment coding for miR‐7b RNA and another fragment coding for a micro‐RNA unrelated to fos mRNA were cloned into vectors, and the recombinant plasmids (designated si‐miR‐7b, samples 3 and 4, and si‐miR‐neg, samples 5 and 6, respectively, in Fig. 1) were introduced into mouse fibroblast cells. Non-transfected cells were used as controls (samples 1 and 2).
Samples 2, 4, and 6, were treated with a chemical PMA that induces transcription of fos gene; samples 1, 3, and 5 were left untreated.
Two hours after PMA treatment protein extracts were prepared from the cultures and were then subjected to Western blot analysis using anti‐Fos (upper panel, Fig. 1 a) and anti-actin (lower panel) antibodies.
RNA was isolated from other cell cultures after 1-hour PMA treatment, separated on a special gel by electrophoresis. A labeled fos cDNA probe was added to hybridize the RNA and imaged. Bands indicate the hybridization of probe to RNA.
1) In your own words explain the purpose of the Western Blot and labeled cDNA probe in this experiment?
2) Why do you think they isolated RNA 1- hour after treatment and protein 2 hours after treatment
3)
The following statements are related to the information presented in the description of the experiment. On the basis of the information given, select:
A : if the statement is supported by the information given;
B : if the statement is contradicted by the information given;
C : if the statement is neither supported nor contradicted by the information given.
a) miR‐7b has a general inhibitory effect on transcription in fibroblasts
b) miR‐7b has a general inhibitory effect on translation in fibroblasts
c) miR‐7b specifically blocks the expression of the Fos protein.
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