Visitor of the Week: Judith Cristobal

Meet Judith Cristobal of the State University of New York at Buffalo! The Filipino citizen is postdoctoral associate in John Richard’s lab and is with us at 2022 course of Expression, Purification & Analysis of Proteins and Protein Complexes. This is second course Judith has taken through our program (the first one was X-Ray Methods in Structural Biology in 2017) and the first she’s taken since the pandemic. She shares that “[although] the advancements in technology greatly helped in making distance learning more accessible during the pandemic, in-person training is still invaluable in providing more hands-on experience on carrying out experiments and more collaborative work with instructors and peers in my field.”

Tell us about your research.
My research focuses on determining the role of a global, substrate-driven enzyme conformational change, in enabling the large rate acceleration of enzymes (GPDH and OMPDC) in catalyzing their respective reactions. My research aims to examine the protein–substrate interactions in enzymes by studying the effect of mutations on the critical amino acid residues that interact with the substrate. We also aim to provide insights into the mechanism for small molecule activation of enzyme activity that is analogous to allosteric activation, while enabling practical uses of chemical rescue in the activation of enzymes for catalysis.

How did you decide to focus on this area/project?
I have always been interested and fascinated by how enzymes carry out their respective biochemical reactions and how enzyme mutations or misregulations lead to disorders. Deeper understanding of the enzyme’s structure, properties, and mechanism is vital for drug discovery and development to treat diseases.

What and/or who is the inspiration behind your scientific journey?
My high school teacher inspired me to pursue a chemistry degree in college. Since then, I have the eagerness and motivation to be a part of the scientific community that works on projects with real-world implications for human health.

Where do you see yourself in five years?
I see myself being a part of a research and development (R&D) team of an industrial/pharmaceutical company that aims to search and develop small drug-like molecules for disease treatment.

What do you love most about being a researcher?
What I love about being a researcher is the chance for the endless pursuit of scientific knowledge. There are always new scientific areas to discover, new fundamental questions to answer, and new open doors to explore.

What drew you to apply to this course?
I have done experiments using traditional methods (IEC; SEC) for purification of GPDH using bacterial system (E. coli). I believe that this program will provide more insights on key approaches such as: a) protein expression in insect cells and mammalian cells; b) protein purification via various affinity methods; and c) protein characterization and analysis such as binding assays and post-translational modifications.

What is your key takeaway from the Course; and how do you plan to apply it to your work?
I learned a lot about protein purification techniques especially on proteins with Tags which I will be applying to purify His-Tag TIM and His-Tag AdK in our lab. The course also invited speakers who discussed cryo-EM and mass spectrometry which are powerful tools that provide valuable structural information for analysis of proteins and protein complexes.

What feedback or advice would you share with someone considering to participate in this course?
I highly recommend this course. There’s a vast array of proteins that needs to be studied and characterized and so taking this course provides course participants with a better understanding of purification protocols and strategies best suited to express and purify their protein of interest.

What’s the most memorable thing that happened during the Course?
The most memorable experience I had in the course is the “chalk-talk“ sessions. Each participant had a chance to discuss his/her research project and ask particular research problem(s) on which they need input. During each session, the instructors and the participants have collaborative conversations to provide the researcher new perspective and approaches on how to deal with his/her respective research challenges.

Is this your first in-person course/workshop since the pandemic? If so, any thoughts you’d like to share?
This is my first in-person course/workshop since the pandemic. Although the advancements in technology greatly helped in making distance learning more accessible during the pandemic, in-person training is still invaluable in providing more hands-on experience on carrying out experiments and more collaborative work with instructors and peers in my field.

What do you like most about your time at CSHL?
I like the quality time spent with the instructors, assistants, and fellow participants during the course breaks. I also enjoyed taking walks at the campus trails and beach front as well as seeing the rich historic architectures all around the campus.

Thank you to Judith for being this week's featured visitor. To meet other featured researchers - and discover the wide range of science that takes part in a CSHL meeting or course - go here.

Images provided by Judith Cristobal

Visitor of the Week: Jessica Rodriguez Rios

Meet Jessica Rodriguez Rios of the University of Puerto Rico - Rio Piedras Campus. In 2016 and as an undergraduate, Jessica joined Dr. José A. Rodríguez-Martínez’s laboratory. Last year, she transitioned into the Ph.D. program and is now a first-year graduate student aspiring to obtain her Ph.D. in biology.  Jessica was recently on campus for her first course, our 2019 course Expression, Purification & Analysis of Proteins & Protein Complexes, and already “can’t wait to come back.”

What are your research interests? What are you working on?
I want to figure out how multi-protein complexes recognize specific sequences of DNA to regulate gene expression. Currently, I am working on determining the DNA binding specificity of transcription factors complexes of GATA4, NKX2-5, and TBX5--all essential for heart development and function.

How did you decide to make this the focus of your research?
I first learned about the molecular aspect of biology in my genetics class and I was intrigued. Subsequently, I joined Dr. José A. Rodríguez-Martínez’s laboratory to do undergraduate research. The Rodríguez-Martínez’s lab mission is to understand how proteins and protein complexes interact with the genome using tools from molecular biology, biophysics and genomic sciences. I knew, without a doubt, that I had chosen the right path. To this day there are many aspects of molecular biology that we don’t understand. The future of molecular biology is promising, and I want to be part of this new generation of researchers that can contribute to a better understanding of molecular mechanisms in biology.

How did your scientific journey begin?
During my senior year as an undergrad, I worked with Prof. Noemí Cintrón tutoring genetics students. Her constant encouragement for me to do research was how I joined Dr. José A. Rodríguez-Martínez’s lab and got my own research project studying the DNA binding properties of transcription factors. This opportunity gave me a new perspective on what it was like to do research. Since then, I have been learning diverse laboratory techniques and sharing my research with the scientific community.  

Was there something specific about the Expression, Purification & Analysis of Proteins & Protein Complexes course that drew you to apply?
For my thesis project, I have to clone and express three human proteins involved in heart development. I applied for the course because I wanted to learn different approaches on protein expression and purification of protein complexes. Our laboratory has been facing some challenges overexpressing soluble proteins, and not having soluble proteins is a major setback and delays downstream experiments. Therefore, for us, it is crucial to have these proteins purified so we can continue our research projects. Also, I wanted to know “best practices” and troubleshooting approaches regarding protein purification.

What and/or how will you apply what you’ve learned from the course to your work?
The instructors gave us a lot of different approaches for protein purification, and I also acquired new perspective on how to approach different steps in the process. Now, I can share it with my colleagues and apply it to future experiments. 

What is your key takeaway from the course?
That every protein is unique, and you might have to use different approaches to purify each protein. Not all of the purification techniques are going to work for a particular protein and sometimes it takes time to optimize your purification process. But that is part of research.

If someone curious in attending this course asked you for feedback or advice on it, what would you tell him/her?
Go for it. The process to apply for the course is very easy and friendly. It was an amazing experience; you won’t regret it. Keep in mind that is very intense, you are going to work in the laboratory from morning to night, but definitely is worth it. You learn a lot about different ways to express and purify proteins with different affinity tags. The course instructors Albert, Michael and Sergei were incredible. They were very helpful, they explained everything and were open to any questions. I highly recommend this course to anyone.

What do you like most about your time at CSHL?
Networking. The team (trainees), we all came from different backgrounds and universities. We got the opportunity to get to know each other as a family. Also, the campus is beautiful, it has a lot of green areas, a beach and beautiful trails where you can take a walk just to explore and think. 

 Jessica received financial support from the National Cancer Institute (NCI) and via a NSF-PR-LSAMP Bridge to Doctorate award. On behalf of Jessica, thank you to the NCI and National Science Foundation for supporting and enabling our young scientists to attend a CSHL course where they expand their skills, knowledge, and network.

Thank you to Jessica for being this week's featured visitor. To meet other featured scientists - and discover the wide range of science that takes part in a CSHL meeting or course - go here.

Repeat Visitor: Tatiana Schnieder

Among the ~9,000 scientists we hosted this year, a number of them participated in multiple meetings and/or courses. Kicking off the 2018 edition of our Repeat Visitor series is Tatiana Schnieder, an assistant professor of clinical neurobiology at Columbia University and an adjunct assistant professor at Hunter College. Tatiana divides her work between two research labs – the neuropathology laboratory of Dr. Andrew Dwork at Columbia University Irvin Medical Center and epigenetics laboratory of Dr. Fatemeh G. Haghighi at Mount Sinai School of Medicine. This year, Tatiana took part in the 83rd CSHL Symposium on Quantitative Biology and joined the ranks of both the CSHL Protein course (Expression, Purification, & Analysis of Proteins & Protein Complexes) and The Genome Access Course (TGAC). Previously, she presented a poster during the Glia in Health & Disease meeting in 2016 as well as attended Blood Brain Barrier that same year. We reached out to Tatiana to chat with her about her experiences in CSHL meetings and courses, and to learn more about what keeps her coming back to campus.

Tell us about your research interests.

My research is focused on the question of brain-periphery interactions in psychiatric disorders. To be more specific, I am currently investigating whether or not the reported relationship between suicide and central and peripheral inflammation is mediated by changes in blood-brain barrier (BBB) permeability.

How did you decide to make this the focus of your research?

During my doctoral training, I investigated prefrontal white matter microglia in individuals who died by suicide; and although I did not find any significant differences in microglial activation between suicides and non-suicides, suicide decedents had significantly higher densities of juxtavascular phagocytic cells in dorsal white matter than diagnostically-matched non-suicide decedents. Interestingly, however, studies of animal models of psychiatric symptoms have attributed maladaptive behavior in rodents to peripheral inflammation and to the influx of peripheral immune cells into the brain parenchyma. These findings and my own findings in postmortem human brains prompted me to investigate the involvement of peripheral immune cells in pathology associated with completed suicide. Further research revealed that non-parenchymal immune cells do not contribute to the increase in perivascular cell density that we observed in suicide; which suggests that resident microglial cells -- rather than non-parenchymal immune cells -- cross-communicate immune responses between the periphery and central nervous system in suicide. Moreover, those who died by suicide had a lower surface area density of microvessels in dorsal white matter -- the same brain region where I found increased densities of perivascular phagocytes. Cumulatively, these findings indicated that changes in the properties of the neurovascular unit could be contributing to pathological changes that lead to suicide. This piqued my interest in the role of BBB in psychiatric disorders. The goal of my current project is to determine if completed suicide is associated with transcriptional, epigenetic, or protein changes in the neurovascular unit.

Expression, Purification & Analysis of Protein & Protein Complexes Class of 2018

What led you to apply for the Protein course and how has it contributed to your work?

I applied to the Protein course to find answers to the challenges I faced during the preparation of my samples for HPLC/mass spectrometry. To assess changes in the expression level of key proteins crucial for BBB function and identify novel differentially expressed proteins, I use HPLC/mass spectrometry proteomics to interrogate protein expression profiles in isolated microvessels. To investigate BBB changes in psychiatric disorders and suicide specifically, I wanted to optimize a microvessel isolation protocol from postmortem frozen human tissue. It was important for me to develop the best method for the extraction of cytosolic, membrane, and transmembrane proteins that seal a gap between the brain endothelial cells. The two-week course gave me a lot of hands-on practice in protein isolation and, through extensive communication with the Instructors and use of the course materials, I was able to come up with the optimal protocol for my project. My main takeaway from the course is that it is imperative to learn as much as possible about your protein/s of interest before you start your experiments. Know your target well because what may work for one protein can be detrimental to another.

How about TGAC? Why did you register for it and what is your key takeaway?

My BBB project involves data that will require knowledge of bioinformatics, and I want to understand what goes into the analyses of the data I produce at the bench. I took TGAC to start learning how to analyze high-throughput data obtained using next-generation sequencing methods and mass spectrometry. As for my key takeaway: Apply what you learn in the course without delay. To acquire a new skill, one needs to practice often – if not daily – therefore it is important to “play” with your data so as to avoid forgetting what you pick up from the course.

The two courses have very different formats: the Protein course consists of hands-on training in a laboratory setting while The Genome Access Course is purely computational. What can you share about the differences or similarities between your experiences in the two courses?

The Genome Access Course Class of 2018 (Spring)

The two courses I took were indeed very different: TGAC was a short two-day course, whereas Protein course was two weeks long and so requires a completely different level of commitment. What is expected of you is also different: TGAC course is mostly lecture-based with some hands-on computer work, and the Protein course is mostly lab-based with lectures sprinkled throughout the day and evening. You cannot really choose to opt out, and by the end of the first week you already feel like you are a part of a team. In fact, working in a team and as a team will be the main mode of learning in a longer course. Irrespective of which course you decide to take, you will be taught by the top experts in the field within a friendly and supportive learning environment, where you will always have the ability to discuss and clarify concepts that may be confusing. It was an amazing experience: I was challenged to step out of my comfort zone, which is how personal and professional growth happens.

The focus of the CSHL Symposium changes annually. This year its topic was “Brains & Behavior: Order & Disorder in the Nervous System,” with an emphasis on neuroscience and related technologies. Besides the overall topic, what attracted you to participate in it?

Once you graduate, it is easy to focus on your specific area of research and, sometimes, you lose sight of the bigger picture. I strive to prevent this from happening to me. As a college professor, I need to stay abreast of the new discoveries in the field of brain research, and attending the Symposium was a perfect way to catch up with the latest findings. It provided a diverse range of topics and speakers from top-notch brain research labs. When this Symposium hones in on this topic again, I would recommend aspiring neuroscientists to attend. If you haven’t yet decided what you want to dedicate your research to or are looking to expand your network, this is a great conference.

Since you’ve now experienced both meeting and course life at CSHL, what differences or similarities did you notice about the two program types?

The courses had a much more focused agenda and required a few prerequisites so you are more likely to meet people who share similar professional or academic experiences as you. The meetings, on the other hand, were eclectic and brought together people from very diverse backgrounds. But in both instances, expect to be surrounded by like-minded people enthusiastic about science and the discoveries it brings.

What did you like most about your meeting and courses this year?

I am very lucky to live a short train ride away from CSHL. Since the first CSHL meeting I attended in 2016 on Glia in Health and Disease, I keep coming back because the quality of the meetings and courses have always been stellar, and the content and organization of the events always meet my expectations. In fact, I am looking forward to attending the Blood Brain Barrier meeting in 2019.

Both the Protein course and TGAC will return to the Laboratory in 2019; and applications are already being accepted. Apply to the Protein course by January 31, 2019 here.

Thank you to Tatiana for sharing with us her experience, and we look forward to having her back at the Laboratory again. To meet other featured scientists - and discover the wide range of science that takes part in a CSHL meeting or course - go here and here.

Visitor of the Week: Archie Reyes

Meet Archie Reyes of the University of Buffalo. The Filipino national is a postdoctoral fellow in John Richard’s lab in the Department of Chemistry. Archie recently made his inaugural trip to CSHL to attend Expression, Purification & Analysis of Proteins & Protein Complexes, and he already has his eyes set on two other courses. 

What are your research interests? What are you working on?
I am interested in enzymes and their modes of action; and studying the general mechanisms on how they provide rate acceleration on several biological processes.

How did you decide to make this the focus of your research? 
While taking my master’s degree, I became really interested in organic reaction mechanisms and protein structure and function and, upon earning my degree, I was sure that I wanted to work in both fields. I realized that the best way to that was to study enzymes – nature’s biological catalysts.

How did your scientific journey begin? 
I have really loved science since I was a kid. It has always been my favorite subject, and I enjoy learning through experiments. My high school chemistry teacher inspired me to start a career in science, specifically in biochemistry.  

Was there something specific about the Expression, Purification & Analysis of Protein & Protein Complexes course that drew you to apply?
There are three reasons why I applied to the course: 1) Learn about other protein expression systems, such as insect and mammalian cells; 2) Be more engaged in proteomics; and 3) Expand my skills in protein purification focusing on protein-affinity tags, as this would help me address our research lab’s need for scaled-up enzyme preparations.

What and/or how will you apply what you've learned from the course to your work? 
I received a lot of help and suggestions from my course instructors and course mates, especially on choosing a more suitable expression system for my enzyme of interest and the appropriate protein-affinity tag to improve the purity of my enzyme preparation.  

What is your key takeaway from the course?
There is so much to learn on purifying and analyzing proteins, and how they interact with other biomolecules. It is important to be familiar with protein biochemisty as it is not just a field of study but also a tool for advancing all life sciences.

How many CSHL courses have you attended?
This is my first time attending a CSHL course, and I am interested in attending either this fall’s X-Ray Methods in Structural Biology or next year’s Cryoelectron Microscopy course.

If someone curious in this course asked you for feedback or advice on it, what would you tell him/her?
CSHL is the best place to learn and obtain new scientific skills in the field of life sciences. If given the chance, I would attend a course from this institution once or twice a year.

What do you like most about your time at CSHL?
I love taking pictures and was able to squeeze in some time to photograph the historically picturesque CSHL! I would love to come back during the fall season to see the institution’s majestic foliage colors.

Archie received a scholarship from the National Cancer Institute (NCI) to cover a portion of his course tuition. On behalf of Archie, thank you to NCI for supporting and enabling our young scientists to attend a CSHL course where they expand their skills, knowledge, and network.

Thank you to Archie for being this week's featured visitor. To meet other featured scientists - and discover the wide range of science that takes part in a CSHL meeting or course - go here.

Visitor of the Week: Mark Dornan

Our Visitor of the Week Series returns with Mark Dornan kicking off the 2017 edition. Mark, a postdoc in Jean DaSilva's Lab at CRCHUM/L'Universite de Montreal (Canada), is on campus for our annual Expression, Purification & Analysis of Proteins & Protein Complexes Course. Read on for more on Mark and his work. 

What are you working on?
I am working on developing new positron-emitting imaging agents to visualize and quantify biochemical processes. Right now, I have two exciting projects with applications in diabetes and prostate cancer.

What is your key takeaway from the Course?
I’ve learned a lot about the fundamental molecular interactions and reactions between the proteins and reagents used for their purification, and how to use this information to make smart choices while generating and improving on procedures.

How many CSHL courses have you attended?
This is my first time attending a course at CSHL. 

What do you like most about your time at CSHL?
CSHL’s harbor-side campus fosters an excellent environment to get to know the instructors and other students. Undoubtedly, my favorite part of attending this CSHL course is the new friends I have made.

Thank you to Mark for being this week's featured visitor. To meet other featured scientists - and discover the wide range of science that takes part in a CSHL meeting or course – go here.