P53 is a tumor suppresser, it controls tumorigenesis via cell apprehension and programmed cell death. In the early 1990s Murine Double Minute 2 ( mdm2 ) was associated with p53 interaction. Oliner et Al. 1993 reported the interaction of the p53 with the N-terminal on the mdm2, caused the suppression of p53 transcriptional activity. Manfredi 2010 showed that in the C-terminal mdm2 consists of a ring finger, giving mdm2 ubiquitin protein lingase ( E3 ) belongingss, doing debasement of p53 through proteasomes. Take together, these observations qualify mdm2 as negative regulator of p53. Donner and Mayo 2001 demonstrated that phosphorylation of mdm2 at serine166 and serine186, encourages mdm2 resettlement from cytol into karyon. In karyon, mdm2 signifiers a complex with p53, later doing its debasement.
What are the major findings of the survey?
Hung et al 2010, reported activation of mdm2 by Akt phosphorylation, and the inauspicious consequence on p53 transactivity. In my sentiment this survey demonstrates and provides farther grounds in vivo, that activated signifier of mdm2, causes suppression of programmed cell death via p53 debasement in mammary epithelial tissue cell ( MEC ) during involution. Over look of mdm2 localised in the nucleus diminishes p53 degrees by mdm2-dependent p53 debasement, therefore ensuing in accelerated tumorigenesis. Sing all of the grounds, it is clear that Akt-mdm2 interaction encourages suppression of p53 mediated programmed cell death, therefore advancing malignant pathophisiology in vivo.
Summarize the experimental attack?
Transgenic mice were generated, utilizing sub-cloning to present complementary DNA encoding mdm2 and mdm2dd into the mouse mammary tumor virus ( MMTV ) booster. This allows the writers to analyze the consequence of mdm2dd on p53 mediated programmed cell death, in comparing to mdm2 and command mice. This transgenic method is considered “ gilded criterion ” . In contrast there is no control over cistron integrating and cistron look. Gene look was measured, at RNA degree utilizing written text PCR, and at protein degree utilizing immunohistochemical staining and immunoblotting. In mensurating transgene look, it allows the writers to find the affect which mdm2dd- p53 relationship has on programmed cell death and tumourigenesis. Increased mdm2dd look leads to detain programmed cell death and an enhanced rate of tumourigenesis due to p53 debasement by mdm2dd. Whole saddle horse and staining techniques were used to detect morphological discrepancies in MECs of control and mdm2dd mice. Apoptosis was measured quantitatively by TUNEL check, at RNA degree by mensurating look of Tweak, LIF and TGFb3 cistrons associated with bring oning programmed cell death. These allow the writer to compare programmed cell death in control mice and mdm2dd mice, and reason how mdm2 affects programmed cell death and its rate by suppressing p53 activity. A t-test, allows the writers to statistically turn out that phosphorylated signifier of mdm2 slowed down programmed cell death and speeded up of tumorigenesis.
How have the writers substantiated the findings?
There is no grounds in proposing that the writers have repeated the experiment to verify the consequences. However their findings are consistent with old probes Hung et Al. 2001. In my sentiment the experiment provides in vivo grounds to the current information, and continues to back up old probes carried in vitro. Application of the T-test is statistically feasible when comparing the degrees of look for mdm2dd and p53, in control and transgenic mice, nevertheless it relies on premises. In replying their hypothesis, the survey uses several methodical attacks. Gene look was measured at RNA degree using written text PCR, and besides at protein level utilizing immunohistochemical staining. Apoptosis was measured utilizing TUNEL check, and besides by mensurating Tweak, LIF and TGFb3 cistrons look associated with programmed cell death in mammary involution. Bitransgenic mice were generated to research the possibility of mdm2 association with other transforming genes in tumorigenesis. The consequences are important in turn outing the aim of the survey, but there is no land interrupting grounds in better understanding Akt-mdm2-p53 tract.
How could the survey be improved?
The transgenic method applied is by and large accepted amongst scientists, one manner to better the method would be to hold a control over cistron integrating and look. The immunohistochemical staining used can do background staining which can interfere with consequence reading. Vosse et al 2007 found that by utilizing HRP labelled polymer no background staining was observed. TUNEL assay can give false positive or false negative consequences. In order to accomplish a high grade of truth, discolorations should be carefully prepared, and more checks performed. The experiment can be improved by repeat to verify the findings, measuring the consequence of mdm2dd in different tissue type.
How has the survey changed the apprehension?
In my sentiment this survey does n’t take bing knowledge any farther. On the other manus it builds upon current cognition by supplying in vivo grounds. It seems that this survey has taken relevant decisions from assorted literature associated with phosphorylation of mdm2 by Akt and its consequence on p53 activity, and tested them in one experiment. These findings were already established prior to this probe. Furthermore it does n’t look in item the molecular mechanisms involved, it merely provides an overview. Writers do non prove a new construct, to take the research to a new degree.
Have the writers achieved their ends?
The writers have addressed the hypothesis suitably. This is reflected by the methodical attack taken. By mensurating the programmed cell death in MECs, the writers prove that mdm2dd leads to detain programmed cell death. Overexpression of mdm2dd in the karyon caused depletion of p53, ensuing in increased rate of tumor development. Conclusively these two measurings reiterate unnatural pathophysiology in vivo. On the other manus, why reference a theory when it has already been proven. In my sentiment there is no grounds that this survey exceeds it nonsubjective, it merely provides in vivo conformation back uping the current literature.
What are the following inquiries?
The first inquiry would be how to forestall mdm2-p53 composite, so that p53 can show its transcriptional activity? How to suppress ubiquitin lingase activity, and phosphorylation of mdm2? Possibly modify mdm2 or p53, so that p53 debasement does n’t happen. See other tracts, binding of p14ARF with mdm2, limits the negative ordinance of p53. Ribosomal protein RPL26 enhances p53 look ( Manfredi 2010 ) . Exploration of these molecular tracts may take to an result which is clinically relevant in the intervention and forecast of chest malignant neoplastic disease.