Identification of Potential Monomeric Transcription Factors in Mycobacterium tuberculosis

Camden Jones Author

08/03/2021 Added

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My UCARE Summer 2021 project on the identification of potential monomeric transcription factors in Mycobacterium tuberculosis

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[00:00:00.840]Hello, my name is Camden Jones. My mentor on this project was Dr. Limei Zhang.
[00:00:05.730]I will be presenting my project from this summer on identification of potential
[00:00:09.390]monomeric transcription factors in mycobacterium, tuberculosis,
[00:00:16.560]microbacterium tuberculosis,
[00:00:18.210]or MTB is the adaptive pathogen that causes the versatile disease tuberculosis.
[00:00:23.610]It affects approximately a third of the population. Every year.
[00:00:27.270]The adaptability of this pathogen is supported by the highly regulated complex
[00:00:31.500]transcriptional regulatory systems.
[00:00:33.600]The pathogen utilizes MTB has been shown to you as approximately 200
[00:00:38.550]transcription factors in order to respond and adapt to hostile host
[00:00:43.110]environments, transcription factors,
[00:00:45.480]regulating transcription by binding to specific sequences of DNA.
[00:00:49.920]The mechanism of these transcription factors can be inferred from its oligomeric
[00:00:53.670]state. And just to give a overview of what oligomeric states are and what I'm
[00:00:58.440]referring to throughout this presentation. I have supplied figure one.
[00:01:03.270]Um, the figure, um,
[00:01:06.390]shows different oligomeric states and what they look like
[00:01:11.490]from a visual perspective.
[00:01:13.740]Um.
[00:01:16.290]As can be seen, uh,
[00:01:18.000]different oligomeric states interact differently with DNA based on the DNA
[00:01:21.900]binding site, which is found.
[00:01:24.060]Here. Um.
[00:01:28.020]If we look at the monomeric transcription factors,
[00:01:30.630]it has a specific binding site that will selectively bind to the monomeric form
[00:01:34.590]of the transcription factor.
[00:01:36.480]Transcription factors can also come in on American states and include more than
[00:01:40.140]one transcription factor. For example,
[00:01:42.510]the next two shapes right here shown in the example are homodimers and
[00:01:46.950]heterodimer transcription factors,
[00:01:49.710]homo dimer transcription factors are transcription factors that bind with
[00:01:53.280]another transcription factor as exactly the same heterodimer
[00:01:58.230]transcription factors are transcription factors that bind to a different
[00:02:03.060]transcription factors. So they're very similar, but just a little bit different.
[00:02:07.650]And then we can also see that it can get a bit bigger as we go.
[00:02:11.460]So here's like another example of a heterodimer and we can go to trimers and
[00:02:16.470]bigger structures such as that.
[00:02:19.380]It is important to note that these are not to be confused as
[00:02:24.480]forming a complex since monomeric transcription factors can also form
[00:02:29.190]complexes with other proteins. And this will be explained.
[00:02:32.190]More in a later slide.
[00:02:34.830]The purpose of this study is focusing on identifying potential monomeric
[00:02:38.280]transcription factors in MTB,
[00:02:40.830]based on a recent study from Fang and their associates A-list of over 160
[00:02:45.030]transcription factors found in MTV were studied and narrowed down to those which
[00:02:48.540]were potentially monomeric figure two.
[00:02:51.120]It gives a more in-depth look at the difference in binding sites for different
[00:02:53.970]odd Americ states.
[00:02:56.280]This figure provides a visual representation of difference between monomeric
[00:03:00.220]transcription factors. Diameric trick description factors.
[00:03:03.940]This figure is included in the original study written by saying,
[00:03:06.460]and their associates uses MOG R, which is here. Um,
[00:03:11.380]as an example for showing this difference as can be seen in the first part of
[00:03:14.800]the figure, when Margaret is functioning as a monomer,
[00:03:17.470]it does not interact with this other transcription factor,
[00:03:20.650]which is also [inaudible] and it can not effectively bind to its binding
[00:03:25.560]region. So transcription is still happening. Now,
[00:03:29.590]if we look at the dimeric state,
[00:03:31.510]it binds with another Mangar transcription factor in this form,
[00:03:35.350]the Maga is able to bind a specific binding site and reduce transcription by
[00:03:39.550]interacting with the Sigma region of RNA polymerase and detach it from its
[00:03:43.360]binding sites.
[00:03:46.030]So why am I interested in identifying potential monomeric transcription factors?
[00:03:50.830]Um,
[00:03:51.970]very little is known about the functional mechanisms of monomeric transcription
[00:03:55.300]factors. So they hold a lot of potential for novel discoveries. Um,
[00:03:59.650]and there is evidence that these transcription factors are vital and interesting
[00:04:02.860]based on known data on the seven web-like proteins.
[00:04:06.510]Why B one through seven found an MTB figure three to fix their
[00:04:10.870]experimental structure by being one in complex,
[00:04:13.600]the Sigma factor a or SIG a this structure was identified by the members imaging
[00:04:18.340]lab. Why B one is a transcription factor found in tuberculosis that has
[00:04:22.300]experimentally been confirmed to be monomeric. As I've mentioned before,
[00:04:26.500]monomeric transcription factors,
[00:04:27.880]conform complexes with other proteins and still maintain your monomeric
[00:04:31.420]oligomeric form. Why do you want us one such protein,
[00:04:35.620]this monomeric transcription factor forms a complex of SIG a,
[00:04:38.770]which is here in order to regulate gene expression in sprinkler system perform
[00:04:43.240]other vital functions and MTB Y B one has also been shown to play a vital role
[00:04:48.190]in cell growth and playing a part in the activation of the dormant form of MTV
[00:04:51.820]in the presence of nitric oxide.
[00:04:55.240]Since tuberculosis affects such a large population, the world, every year,
[00:04:58.720]we need new ways to effectively treat this deadly disease,
[00:05:01.690]learn any more about the mechanisms of these monomeric transcription factors can
[00:05:04.930]provide new opportunities to formulate new drugs and therapies to treat
[00:05:08.350]tuberculosis more insight into these monomeric transcription factors can also
[00:05:12.400]contribute to understanding how MTV functions and how it is able to adapt so
[00:05:16.450]well in host stress environments and where is to generate a list of potential
[00:05:20.680]monomeric transcription factors, and then test on dimeric steaks.
[00:05:23.800]Several methods are utilized.
[00:05:26.140]These include included several methods of sequence analysis, PCR, cloning,
[00:05:30.670]protein overexpression,
[00:05:31.660]and pull down and SES for the purposes of this presentation of my UCare project.
[00:05:35.530]I'll be focusing on the methods of sequence analysis in order to describe
[00:05:40.390]him as a process and identify these potential monomeric transcription factors.
[00:05:44.110]I'll be using the transcription factor RVs zero, zero to three, as an example,
[00:05:48.970]databases such as micro browser, you know,
[00:05:50.830]pro and blast for use the analyze the sequences of each transcription factor and
[00:05:54.610]to determine their possible functions, structure, prediction tools,
[00:05:58.220]such as Swiss model and Kymera reuse to visualize and determine the putative
[00:06:01.940]oligomeric state of the transcription factors.
[00:06:04.910]Micro browser provided information information such as the located shin of the
[00:06:09.590]gene and MTV's genome and its possible function.
[00:06:12.590]And this will be shown in figure four blast shows matches the XRD family type
[00:06:17.180]transcription regulators and Swiss model Fenn,
[00:06:20.000]a homology model that was monomeric. This will be shown in figure five.
[00:06:24.200]Tamera was then used to help visualize the hemology models found for the gene
[00:06:29.150]when necessary and was used to identify the potential all Americ state.
[00:06:33.560]Thank you for shows the corresponding Michael browser page for RV 0 2 3.
[00:06:38.270]This page shows the RV 0 0 2 3 is located upstream from Y V five.
[00:06:43.580]Um,
[00:06:44.810]this page also highlights that the transcription factor is possibly involved in
[00:06:49.310]a transcriptional mechanism. From this page,
[00:06:51.500]I was able to find hemology models from Swiss model that correspond to RBA
[00:06:55.060]0 2, 3,
[00:06:57.500]figure five shows the corresponding Swiss model page with the homology models
[00:07:01.370]found in the make of other page as can be seen.
[00:07:03.860]The homology models are attached our monomeric.
[00:07:07.850]This can be an indication that RV's year zero to three is also potentially
[00:07:11.660]monomeric
[00:07:13.850]from the list of over 160 transcription factors in MTV from things study,
[00:07:17.900]I was able to narrow it down to 15, potentially monument,
[00:07:21.140]potentially monomeric transcription factors,
[00:07:23.600]things study had 32 potentially monomeric TransPerfect transcription factors
[00:07:27.950]listed.
[00:07:28.880]I was then able to further narrow down the list of seven monomeric transcription
[00:07:32.390]factors that I'm interested in pursuing in my future work.
[00:07:35.720]I'm only interested in these transcription factors that show evidence of
[00:07:39.080]actually being involved in gene regulation. For example, um,
[00:07:42.950]if one showed the possible function of being an enzyme that didn't directly
[00:07:47.810]involve gene regulation,
[00:07:49.790]I did not include it in my list due to certain safety precautions,
[00:07:53.900]other genes that may have been shown to be potential toxin antitoxin systems.
[00:07:59.240]Um, I admitted those from my list as well.
[00:08:02.660]And from my sequence analysis narrowed down list includes RVs 0 0 2 3 are
[00:08:07.360][inaudible] RV 1359 R V
[00:08:11.450]1830 RV,
[00:08:12.980]1931 C RVT 9 0 1 C and R B 3 8 4 0.
[00:08:19.550]My most current work on this project includes preliminary work in the lab with
[00:08:23.180]the help of Dr. Jang and other lab members.
[00:08:24.980]I need to successfully purify plasmids containing the construct for RV 1830 and
[00:08:29.480]RV 2 9 0 1 seat.
[00:08:31.610]I'm currently testing the expression level of the RV 1830 construct with
[00:08:35.030]small-scale overexpression in small scale purification for my future.
[00:08:39.880]Sure work.
[00:08:40.300]I plan on continuing this project for my student thesis and for feature you care
[00:08:43.870]projects. After experimenting with RV 1830 and refining my protocols,
[00:08:48.280]I will move on to test the other transcription factors in my narrow down list,
[00:08:51.910]using the methods such as PCR, cloning, protein over and pull down states.
[00:08:56.910]These methods in the data produced will help me determine whether the list of
[00:08:59.970]genes are in fact monomeric transcription factors or not.
[00:09:03.750]This work was supported by the undergraduate creative activities and research
[00:09:07.050]experience program. And I want to give special thanks to Dr.
[00:09:11.070][inaudible] Daisy, Giza, Beltran,
[00:09:14.560]and Taiwan for guiding me through this project and meeting amazing mentors.
[00:09:17.910]Thank you for your time.

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Identification of Potential Monomeric Transcription Factors in Mycobacterium tuberculosis

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