39 thoughts on “Ionizing-radiation Toxicogenomics”

1. Human exposure to radiation is a common topic popping up in the scientific community today. Radiation exposure is responsible for cellular and DNA damage in human beings. Radiation is all over the place today including the working environment. According to Denis A. Smirnov, et al., the effect of radiation on gene expression differs in individuals. These scientists have performed genetic tests in order to identify the chromosomal regions that influence the expression of several genes due to radiation exposure. The researchers used microarrays to measure the expression levels. They used immortalized B cells for this experiment because they are sensitive to radiation. Data was collected at baseline, 2 hours and 6 hours after the B cells were exposed to radiation. There are two different regulatory factors mentioned in this paper, the cis and trans regulatory factors. The different between cis and trans linkages is that a cis linkage regulates the expression of genes that are located on the same molecule and trans linkages regulate the expression of genes located on a different molecule.

   An example of a cis regulator is PHLDA3. This gene is also known as the Pleckstrin Homology-like Domain, Family A, Member 3 gene. PHLDA3 is a tumor suppressor gene that inhibits AKT signaling. It contains 2 exons. PHLDA3 is required for tumor suppressor p53-dependent apoptosis.

   An example of a trans regulator is LCP2. This gene is also known as the Lymphocyte Cytosolic protein 2. LCP2 encodes a signal transducing adaptor protein. Evidence has shown that this gene plays a strong role in promoting T cell development and activation in mice.

   • Its really interesting to see even though the trans-acting regulators were found to be dominating that in your case, PHLDA3 as you mentioned to be a tumor suppressor is actually a cis-regulator. I would think it would be the latter since it is required for apoptosis. In the article is was founded that BAX which was trans-acting was actually a trans-acting regulator. I would think it would correlate with this evidence. Also, interestingly enough it seems since LCP2 is more of an immunological response which. I don’t know about you, but it seems a bit obscure to me because although cis-regualators were less than 1% IR induced expression it seems to play a pretty significant role in my eyes.

     • But what is the difference between a cis and trans in this case? Can you think of a physical example of each?

2. The article is about a research done using radiation as a drug therapy, it is divided into the radiation tolerance from sensitivity to insensitivity to the influence of radiation. Change in gene expression is based on the damage caused by the radiation. Radiation kills the cells by destroying the cell’s genetic material. Person’s genome determined differential responses to the therapy, and the most common response to radiation is apoptosis, which is programmed cell death.
   Researchers conducted this experiment using immortalized B cells and using technologies like microarray that identifies the expression of genes after being exposed to radiation in two hour and six hour intervals thinning ten thousand genes narrowing them down to around three thousand genes. They also used SAGE to carry out genome wide linkage analysis for the 2 and 6-hour radiation expression phenotypes as well as the technology Chilibot a program that scans through papers finding word associations and identifies the relationship between certain transcription factors and specific genes.
   The results showed the variation in gene expression response to ionizing radiation. They found TP53BP2 on chromosome 1 and is a regulator of BAX 19. The paper was much interesting to read since I already had knowledge on microarray; it was interesting to read the use of microarray for existent works.

   • TP53BP2 is an example of a trans-regulatory factory. It regulates BAX is involved in apoptosis when exposed to radiation. What is an example of a cis- regulatory factor?

3. Researchers (Smirnov et. al.) used several techniques and ironically enough microarray analysis was used to measure the expression levels of genes in irradiated immortalized B cells especially from members of CEPH. The data was compared at both 2 hours and 6 hours after exposure to (10 Gy) of ionizing radiation. Expression data was obtained for cell lines derived from 2 parents and 8 children per each family. Researchers wanted to focus on ‘ionizing radiation responsive’ genes that showed at least a 1.5-fold change in gene expression. The ratio of expression level was calculated and compared to at 2hours and 6 hours exposures. Remarkable data was obtained (4,600 single nucleotide polymorphism) markers were obtained showing significant linkage. Furthermore, family-based association analysis was done to confirm the linkage finding s and obtain more accurate results. This concluded in testing each cis-regulated gene for association by (QTDT) a type of test using SNP markers. This test contributed to the importance of cis and trans- acting regulators. Thus, after 2 hours IR certain phenotypes were prominent and after 6 hours IR different phenotypes were prominent. In comparison, the expression levels are defined as regulated by the trans-acting regulators. This was seen via “trans peaks” thus, discovering BAX that plays a role in apoptosis. Conclusively, this facilitated in determining that IR-induced expression phenotypes are trans-acting factors. Trans regulation provides cells with multiple mechanisms to mount response to different types of stressors and permits coordinated responses since it also influences expression of several genes.

   • I’m not sure if you picked up on this also, but did you wonder why the researchers chose such specific time slots to monitor the radiation effects on the B cells? I was wondering if they chose baseline, 2 hours, and 6 hours for a specific reason or if it was a random selection. What do you think?

     • You could always ask the author: vcheung@mail.med.upenn.edu

     • The reason they probably picked 2hours and 6 hours is to see the effect of radiation lengthwise and compare the results between those cells. Longer someone is exposed to radiation the cells would be damaged more.

4. Radiation is a huge toxicological issue because radiation is known to cause changes in gene expression that are often harmful for the organism exposed to radiation. Radiation is also used in diagnostic and therapeutic medical technologies, so knowing what radiation does to cells is very important. Researchers used microarray analyses to test the change in gene expression of cells exposed to radiation. The researchers chose immortalized B cells to achieve this. This is because these particular cells are highly sensitive to radiation-induced gene expression changes. They measured gene expression changes from the baseline expression levels at 2 hours after radiation exposure and 6 hours after radiation exposure. They examined the effects on cis- and trans-regulatory factors. A cis-regulatory factor is located on the same part of the genome as the genes it regulates, while a trans-regulatory factor is not located near the genes it regulates. The researchers showed that, in most cases, trans-regulated genes were the ones that were most affected by the radiation exposure.

   An example of a cis-regulatory factor from the paper is PHLDA3. This is a gene involved in apoptosis that is regulated by p53. It represses AKT1 activity by preventing AKT1 from binding to the cell membrane, and this action promotes apoptosis. This gene is a tumor suppressor.
   An example of a trans-regulatory factor from the paper is TP53BP2, which is also a tumor suppressor gene that interacts with p53 protein to promote apoptosis when the cell is under stress.

   • PHLDA3 example for cis-regulatory factor is same as p53 target gene which encodes a PH domain protein. P53 and Akt negatively regulate each other to balance survival and death signals within a cell.

5. The exposure to Radiation affects individuals differently. Radiation causes damage to the DNA. Scientists used Microarray analysis to measure the expression level of cells in immortalized B cells. The researchers chose B cells because these cells are highly sensitive to radiation. Researchers observed the mechanism of cis and trans regulatory factors. The data obtained were after two and six hours of exposure. The chromosome location was done by a genome scan. A computer program called S.A.G. E was used to observe the genome linkages for the 3,280 genes. A cis regulatory factor is located on the same gene it expresses while trans regulatory factor is not located on the same gene. The cis regulatory factor and trans regulatory factor analysis differed. The cis regulatory factor had only 9 which is less than 1 percent and were regulated after two hours of exposure and after 6 hours only 12 were regulated which is less than 1 percent. While the remaining phenotypes were regulated by trans regulated factors. This provides evidence that the trans regulated genes were impacted the most by the exposure of radiation.

   An example of a cis regulator would be PHLDA3. This particular gene induces apoptosis and is regulated by p53. It represses AKT1 by preventing it to bind to the membrane. It is also associated with p53 dependent apoptosis by inhibiting the AKT1 activity. It controls transcription by negative regulation of AKT1 it is also , known as a tumor suppressor.

   An example of a trans regulator would be TP53BP2, it plays a significant role in regulation of apoptosis, It is also known as a tumor suppressor gene. This gene is located on chromosome 1.

   • Does this experiment show that PHLDA3 is important in to not become suppressed because being suppressed may result in cancer?

6. The genetic analysis of radiation-induced changes in the human gene expression was carried using different methods to analyse and verify the researchers findings. These series of experiments was done using immortalized B cells from the same family so that the genetic information and location of genes would be consistent throughout all cells. The first Step in their experiment was the genetic mapping of regulatory elements, which was done by using microarrays to measure the expression levels of genes. The cells were grown, irradiated, then harvested after 2 and 6 hours after exposure. RNA was then extracted and hybridized onto arrays. The next test they ran was for cytotoxicity. cells were harvested based on their over or under expression of JUN, then irradiated and observed over the coarse of 24 hours to study how resistant they were to the radiation. The next step was the analyses of linkage, which was done by a program SiPAL. SIPAL determines linkage based on phenotypic differences between siblings. The then proceeded to observe the hotspots of the genome by dividing the autosomal genome into windows and monitored them after 2 and 6 hours after being irradiated. a hotspot is a region of the genom that probably contains regulators the influence the expression of genes. the last step in their experiment was the knockdown of candidate regulators. they did this by transfecting candidate regulators and non-target controls with siRNAs to study the effects.

   Cis-Regulatory factor regulates genes on the same chromosome, in this experiment the confined “cis-regulators” to be within 5 mega bases of target gene. An example of a cis-Regulatory factor from the paper would be PHLDA3 it is a factor in increasing cytotoxicity and capase levels.

   Trans-Regulatory factors regulates genes on different chromosomes. an Example of a tran-regulatory factor would be CD44 a cell surface receptor that regulates TNFSFR.

   • How exactly does PHLDA3 function to increase cytotoxicity and capase levels in the cell?

7. Although many people may have been paid attention to the influences of radiation exposure, few are aware of radiation-induced changes in our body, which is in human gene expression. However, researcher, such as Denis A. Smirnov, is able to examine this issue by obtaining a microarray so that the expression levels of genes disease cell, irradiated immortalized B cells from members of 15 Centre d’ Etude du Polymorphisme Humain Utah pedigrees, can be measured. And Smirnov’s group began with gene expression response to radiation because cellular responses rely on changes in gene expression. By examining genetic linkage and association analyses, Smirnov’s group is able to identify DNA variants that influence the expression levels of genes in irradiated cells. And the result turns out to be that nearly all of the phenotypes of cells at the baseline are regulated by trans-acting factors. One of the most identified features of trans-acting regulator is that they are transcription factors, such as RB1, HIVEP2 and UBA52. In addition, another way to differentiate cis- acting regulator and trans- acting regulator is by its target gene landscape. For instance, cis-regulators are considered to be those that were mapped within 5 mega bases of the target gene, such as CP110 at location 16p13-p12 (Fig.2). Thus, by having a genome-wide perspective and using all these advanced biotechnology, the mystery of human gene expression involved in radiation exposure can be figured out. Moreover, our understanding in gene regulation will develop and expand more eventually.

   • What does RB1, HIVEP2 and UBA52 regulate in the human body?

8. In Smirnov’s , et al., article called Genetic Analysis of Radiation- Induced Changes in Human Gene expression, they try to identify the genes that may serve as potential markers of radiation sensitivity. They go about doing this by using microarrays to explore gene expression after cells are exposed to radiation. The cells that they use are immortalized B cells from 15 memebers of the Utah pedigree that donated their DNA to the CEPH. They decided to use these cells, because they are very sensitive to radiation so they will show changes in gene expression to even minute amounts of radiation.
   Cis- regulatory and trans- regulatory factors were also mentioned to further differentiate the genes that were affected by radiation in the experiment. Cis-regulatory is when a gene is present on the same chromosome as the gene it regulates, while trans-regulatory regulates genes distant from the gene that it was transcribed. In the experiment any gene mapped within five megaabases of the target gene was considered cis-regulatory, while all other significant linkage findings were considered trans- regulatory. Most of the genes that were affected were found to be trans-regulatory, while very few were found to be cis-regulatory.
   One example of of cis-regulatory is CP110, which has the protein involved in centrosome duplicaition. In the experiment the gene lead to unschdeuled centrosome duplictation, chromosomal instability and cell death after the six hour irradiation mark.
   An example of trans- regulatory is the expression in ZNF493, which is a gene that is involved in transcriptional regulation. Only after 2 hours after irradiation this gene showed changes in gene expression.

   • So which one is the regulator? and what will regulate the regulator?

   • Nice job discussing the procedure of the paper. I think that if you mentioned more about ZNF493, and all of its functions how it effects the trans-regulatory genes it would further improve your paper.

9. In todays modern world everyone at some point of the day is exposed to radiation. The experiment conducted on the genetic analysis of radiation-induced changes in
   human gene expression lead to the conclusion that radiation effects are different depending on the person that is affected. In this experiment immortalized b cells were studied two hours and six hours after exposure to ionization radiation. Immortalized b cells are cells that do not stop division at any point and can cause tumors in the body or cancer. Micro array analysis was used to gather the information on the cells via mRNA. These cells were studied and a gene list was comprised with the results separated by cis and trans. Cis meaning the same molecule and trans meaning separate. The guideline used for determining whether the results were cis or trans was that anything that gave a megabase of five was considered cis and anything other was trans. These genes released regulators which were important in the study as the cells releasing the regulator Bax which led to apoptosis were an important factor in this experiment. The conclusion of this experiment was that radiation affect everyone however it had its own different affect on different people. This experiment was conducted using cells for 30 people of 15 different pedigrees which mean that for every person from the same pedigree they had the same susceptibility for certain diseases based on their genetics.

   • I think this shows a well understanding and good summary of the text the only problem i see is the way you described the difference between cis and trans. The 5 mega-bases refers to the base pairs in their sequence and just mean that of that DNA on that chromosome they lie only 5 mega bases away from each other meaning they are extremely close when cis and noticeably further away for trsnd

10. Radiation is an important toxicological issue because we are exposed to radiation through the environment. In addition, radiation is used in diagnostic and therapeutic medical technologies. However, radiation is can induce DNA damage in cells, causing changes in gene expression. In response to radiation cells mount complex responses that depend on changes in gene expression. It is known that individuals vary in their in their sensitivity to radiation and this variability is observed in the gene expression level.
    Researchers have used microarray analyses to test the change in the gene expression level of cells that have been exposed to radiation. First they radiated immortalized B cells from members of 15 Centre d’ Etude de Polymorphisme Humain (CEPH) Utah pedigree. The reason researchers chose cells from a pedigree is because of the close and known relationship among individuals. In addition, immortalized B cells were chosen because they constantly divide. After the cells were exposed to radiation, mRNA was extracted to perform the microarray analysis. From the analysis, data were collected for cells at baseline and at 2 and 6 hours after exposure to 10 Gy of ionizing radiation. The data provided researchers wit a list of genes that they separated into cis and trans.
    The data showed that nearly all of the regulators act in trans to influence the expression of their target gene and very few are cis-acting regulators. The difference between cis and trans is that cis-regulatory factors is that cis regulatory factors regulates the expression of genes located on the same chromosome while trans-regulatory factors regulate the expression of genes located on different chromosomes. A regulator is determined as either cis or trans on the basis that cis-regulators are those that were mapped within 5 mega bases (Mb) of the target gene and any other linkage is considered trans.
    An example of a cis-regulatory factor is CP110. CP110 encodes for a protein involved with centrosome duplication. The disruption of CP110 leads to unscheduled centrosome duplication, radiation- induced chromosomal instability, and cell death. An example of a trans-regulatory factor is TP53BP2. TP53BP2 is located on chromosome 1 and a known regulator of BAX. The radiation-induced expression level of BAX has a role in apoptosis.

    • http://omim.org/entry/246530

      • ignore link above

    • I think you have a very good understanding of the concepts involved in this study. What was the actually process the researchers had to do to get this information?

11. Every person is exposed to radiation at some point of their life, making radiation an important toxicological issue. The paper Genetic Analysis of Radiation-Induced Changed in Human Gene expression by Smirnov et. Al. identifies some genes that went through radiation-induced changes in human gene expression. Researchers did this by taking human immortalized B cells from the same pedigree and exposing it to radiation. They used this type of cell because it doesn’t stop dividing and since they are from the same pedigree, researchers know the relationships and variability that accompany these cells. Researchers exposed the cells to radiation in different doses (2 hours and 6 hours). Then they extracted the mRNA from the cells and did a microarray on the cDNA that was made from the mRNA. They analyzed the microarray and came up with a list of genes that were expressed differently. With this list they analyzed the genes even further using different databases. Cis-regulatory factors are factors that regulate gene expression that are on the same molecule, in this experiment within 5 megabases. Trans-regulatory factors are factors that regulate gene expression that are not on the same molecule, in this experiment over 5 megabases.
    An example of a cis regulator is PHLDA3, the Pleckstrin Homology-like Domain, Family A, and Member 3. This gene is required for tumor suppressor p53-dependent apoptosis. It represses AKT1 activity by preventing AKT1 from binding to the cell membrane.
    An example of a trans-regulatory factor is TP53BP2. IT is a tumor suppressor gene that interacts with p53 protein to promote apoptosis when the cell is under stress.

    • Your response was very detailed and articulate. I liked how you were very particular in explaining exactly what was done for the experiment. The part that seemed to really bring this response into perspective would have to be the introductory sentence which shows importance to the experiment and to your response.

12. The researchers did this by getting a pedigree of human cells turning them into immortalized B cells and irradiating them. Afterwards the scientist performed a microarray analysis and used several programs to produce a computational analysis of genes. Cis regulatory factors are within 5 megabases of each other and trans-regulatory factors are out side of 5 megabases. An example of a cis regulatory factor is FAM57A and an example of a trans-regulatory factor is EDG1. CP1110 encodes a protein for centrosome duplication. FAM57A which regulates the cell cycle and apoptosis.

    • This is a good response because it answers all of the questions being asked but I think that you could go more in depth. What kinds of programs did they use for the microarray? How long did they expose the cells to the radiation? If you explained a little more I think tha tthis response could be of better quality.

13. Radiation is a very dangerous substance that a group of researchers are looking into. Radiation can be found in medical settings in the form of x-rays, or even in the environment. Cells must go through a gene expression change to accommodate for the radiation poisoning. First, the group of researchers took a group of human immortalized B cells. They knew that there was a relationship between the people that the cells came from. This allowed them to use a pedigree which could tell if there has been any disease or complications in the cell line. After they obtained the cells, the researches radiated them two different lengths of time: two hours or six hours. They then took the mRNA out of the cells and completed a microarray. They found that there were two different categories of regulatory factors: cis and trans. The cis regulatory factors are located on the same molecule within five mega-bases away from the DNA sequence being examined. An example of this is the DNA sequence right next to the gene that is about to be regulated, that is within five mega-bases away. Trans regulatory factors are more than five mega-bases away from the DNA sequence being examined. An example of a trans regulatory factor is a transcription factor.

    • Also, B cells are immortalized and sensitive to radiation. I like your post you write very clearly for all readers to understand!

14. Researchers used microarray of irradiated ionized B cells to study gene expression after exposure to radiation. The B cells were immortalized, which means they regenerate quickly and don’t die, giving the researchers a multitude of cells to work on. B cells are also sensitive to radiation, so the effects of the radiation are clearly expressed.
    The difference between a cis and trans regulatory factor is the distance of one regulatory factor and the other. Cis regulatory factors are within 5 megabases of each other. Trans regulatory factors are outside of 5 megabases of each other.
    An example of a cis regulatory factor is PHLDA3, a gene involved in apoptosis regulated by p53. It prevents AKT1 from binding to cell membrane, promoting apoptosis.
    An example of a trans regulatory factor is TP53BP2, a tumor suppressor gene that also interacts with p53 to promote apoptosis.

    • That was an interesting factor chosen. I am wondering though what exactly the tumor-suppressor gene p 53 isn’t involved in? So far you have showed a great coonnection to this p 53 and one of the genes I chose last paper discusion I posted.

15. Radiation causes many mutations in gene expression. Though radiation is extremely harmful we run into exposure everyday. For example X-rays, Cancer treatments, and many tests for diseases are conducted using radiation. In this article researchers tested gene expression in exposed cells compared to non exposed cells using Microarray Technology. The cells that were tested were immortalized B-Cells, due to their high sensitivity in radiation-induced gene expression. The cis- and trans-regulatory factors were examined. The cells most effected were the trans-regulatory genes. PHLDA3 is an example of a cis-regulatory factor. This is involved in preventing the AKT1 from binding to the cell membrane, promoting apoptosis. This promotes apoptosis. LCP2 is an example of a trans-regulator which promotes Tcell development.

16. In the beginning the microarray experiments were of big help to start location gene expression profiles. The researchers were identifying the genes from there with the S.A.G.E. statistical analysis of genetic epidemiology which allowed specific and various studying of multiple genes. An example of Cis- regulatory is the CP-110 that is GRAPHED on page 2 of this paper. This functions on chromosome 16 to show any irradiation exposure in the molecule gene’s expression.
    The CP-110 is known for the association of Kif24 kinesin to remodel organelles in the cell within centrioles.

17. Regarding the ASL: March 4, 2012 – March 10, 2012
    This academic servie learning comes from taking the real time out of a busy normal school day and being able to replace the that with the community service. The material of Toxicogenomics can be overwhelming and so be able to teach such can only enhance one’s own personal learning, engagement, and for the other’s as well.
    I had observed that the junior high school students were well-versed and were knowledgeable of the information and the material being covered in class that day.
    Especially, my student was so thorough and even enthusiastic about the entire lab experiment. The actual lab manual designated for the younger group was amazingly easy to use and read for the students. Some students decided to take home a copy and allowed them to recall details while learning more of the process.
    From Lab 7, done personally, the insight gained was profound as to how this specific topic of E.L.I.S.A is used in everyday and home held products or activities or tests. The analytical understanding of the E.L.I.S.A. used examples from home tests for pregnancy, the presences of infection from viral or bacterial.
    Now, in completing Lab 8 with the middle school students, the teaching experienced entailed for me to have my pre-knowledge and understanding in order to effectively communicate in more simpler and pedantic terms, too. The transition from the student to teacher was simple having a trial of the lab done under my belt.
    Genuinely, I felt that the there was a way more we could do as a student body for the Junior High kids. The suggestions to be made would be to create more activities; color-in pages, or a follow up visit with sharing or results shown type of day.
    Overall, the day was a great ASL experience for myself and my student in learning the E.L.I.S.A.

18. Regarding the s i RNA therapy working: March 27, 2012 – April 3, 2012
    SiRNA therapies are a future technology which aims at gene silencing by RNA interference. SiRNA uses a specific mRNA which targets degradation for the inhibition of the synthesis of the encoded proteins. SiRNA therapy works by making a change in the base, sugar, or backbone structure of the RNA of the cell. Thus causing a mutation in the DNA (gene) transcribed causing silencing of the gene. When you add a synthetic SiRNA molecule to a cell the affect is gene silencing. At the molecular level this is done by ensuring a safe delivery method (not being affected or attacked by innate immunity) and having the SiRNA up taken by the cell. After uptake the siRNA must escape from endosomes or lysosomes and has to avoid compartmentalization. At the sight of action (nucleus) must be available and then gene silencing activity is executed. The information that should be known about the gene target is what the affect would be if the gene is silenced. It is also important to know if there are similar genes that would also be silenced and what the affect would be if it is silenced. The safety of siRNA is still questionable and is still being continued to be studied. Personally, if this has a higher success rate then its a more effective and should become more widely known therapy universally. So far, I noticed a correlation between this and the other papers of How Complex is Life and the Tackling Asthma in Childhood.

19. Regarding the Many, many genes in Toxicogenomics:
    Microarrays are the new useful and easy to use/perform technological advances. Growing after the late northern Blot tests were used, the take over from microarrays express the gene signatures and profiles from two trials, controls DNA and the samples DNA, cDNA and sDNA. The TPA, 12-O-tetradecanoylphorbol-13-acetate, common gene name TTMP. The use of DNA microarrays, according to the paper, for intervening the gene expression patterns of cells exposed to the outside toxicants and the small molecules are the products that could yield insight to the drug/chemical – induced toxicity or cancer. So now in reference to this protein that is a tumor promoter, a gene expression of change in cells that were exposed to the anti-cancer agent such as TPA. In any species many many genes affect others and proteins the same. So in a microarray a signature of the specific genes from normal or exposed cells will release their profile either by color or clustering. In Lab 9: Microarray Experiment was conducted and compares to this situation easily. Understanding the Microarray is necessary to better grasp this experiment’s point. The main idea is to use such simple methods to determine thorough quick and effective results.

Comments are closed.