July 2021: Kalderon Lab

Background:

What is the main focus of your lab?

Please see https://kalderonlab.biology.columbia.edu/

We aim to understand how Drosophila Follicle Stem Cells (FSCs) fulfill their function as stem cells during normal physiology. This involves defining where FSCs are, what they do (divide, change location, differentiate into either an Escort Cell (EC) or a Follicle Cell (FC)), and how these behaviors are instructed by positional signals (relayed by Wnt, JAK-STAT, Hh and other ligands). If the scientific community can understand the cellular and molecular organization and logic of a variety of stem cell paradigms there would be a solid foundation for better understanding (a) how to manipulate stem cell function therapeutically and (b) how specific types of mutation can initiate  cancers. FSCs present one such paradigm; they have a number of favorable characteristics, are already quite well understood and have numerous similarities in organization to mouse gut crypt stem cells. FSC studies have clarified the different roles played by key niche signals and the selective advantage conferred on a stem cell through increased proliferation rate for any paradigm where stem cell division and differentiation are not coupled events. The lab also examines the mechanism of Hedgehog (Hh) signal transduction in wing disc cells in a separate project.

How long have you had your lab? When did you join Columbia University?

I joined Columbia in 1988, worked initially on the functions of Protein Kinase A (PKA) in flies, leading to discovery of involvement in Hedgehog (Hh) signaling (1995). Studying Hh signaling in oogenesis, to supplement embryo and wing disc studies, led to the realization that Hh regulates FSC behavior (2001), and a decision to study FSCs more generally.

How big is your lab currently?

One Research Associate, two PhD students, two Research Assistants, one undergraduate.

Where is your lab located?

1013 Fairchild, Morningside campus

Current affairs:

What are the most exciting projects/directions in the lab at this moment?

We hope to understand the cell biology and molecular mechanism of an FSC becoming an FC through identifying molecules (likely including adhesion molecules) that affect this propensity and respond to the external signals that regulate this behavior, and perhaps by visualizing the process better during live imaging using GRASP techniques to see cell contacts.

We also hope to be able to obtain significant insight into how external signals regulate the rate of FSC proliferation, by continued genetic studies and use of cell cycle reporters, to discern mediators and the cell cycle transitions affected.

We have studied individual FSCs extensively to understand cell autonomous responses to environmental signals. We would also like to understand how the behavior of a community of stem cells differs from the simple sum of responses of individual stem cells. In effect, how does the signaling environment and other interactive parameters (cell contacts, for example) change in response to perturbations in order to maintain function of the stem cell community? We are exploring this by imposing global perturbations affecting cell cycling and key graded extracellular signals.

What are the biggest accomplishments that your lab recently had?

Several years of work prior to 2017 led to a publication that completely revised our understanding of FSC behavior (large number of mobile stem cells supported by population asymmetry, supplying both ECs and FCs rather than just two FSCs supplying only FCs through single-cell asymmetry). These revisions were important not only because the old model of two decades was incorrect on many counts, but also because they allowed measurement and understanding of a far richer set of individual stem cell behaviors. The long and continuing experience of challenging dogma has, however, generally been difficult and frustrating, exposing weaknesses of the scientific system to which we aspire. It also has revealed an apparently common lack of appreciation of some key, universally applicable concepts for defining stem cell behavior from lineage analysis. I hope to capture those insights in a review or opinion piece.

More recently, thorough analysis of the consequences of altering Wnt and JAK-STAT signaling (which collaborate with Hh, PI3K, BMP, integrin and Hippo/Yki pathways) has produced a self-consistent framework for understanding how the FSC domain and differentiation into two different cell types at each domain boundary are organized by graded extracellular signals. Altogether, our studies have shown that FSC behavior is very fluid, apparently responding rapidly to position-dependent signals and with little evidence of rigid boundaries, remembered behaviors or sharp transitions in gene expression patterns.

Technology:

What are the model systems that your lab is using? 

Drosophila melanogaster- we have large numbers of stocks for manipulating the activity of various signaling pathways and their potential mediators, cell cycle reporters and various lineage analysis tools.

What are the key techniques that your lab is using? Are you open to training scientists from other labs?

The majority of our work involves standard tools for Drosophila genetic analyses, together with relevant dissection, staining and imaging. We also have live imaging capability using a chamber made with the the help of former members of Gordana Vunjak-Novokovic’s lab. We are open to training.

What facilities or equipment does your absolutely lab rely upon? Do you use CSCI cores?

We do not use any CSCI cores partly because we are on a different campus. Here, our crucial “core” is the Departmental (Biological Sciences) confocal facility.

Who shall be contacted with questions about equipment, resources and training?

Dan Kalderon

Training:

What's your best approach to mentoring trainees in the lab?

The most important mentoring objective that I actually execute robustly (many potentially beneficial approaches are acknowledged but not consistently put into practice) is rigorous appraisal of all aspects of data (experimental design, veracity, flaws, logic of interpretation and limitations) through repeated discussion, new or repeated experiments, and re-organization of presentation or processing. I believe the difficulties in designing adequate tests and interpreting results are often underestimated by experimenters and reviewers, leading to several published false trails and beliefs. Precise thinking and arguments, being open-minded, skeptical and continually considering alternative scenarios or arguments are key traits to develop through persistent practice. Raw data are absolutely crucial and are often the subject of discussion. However, there is no adequate substitute for the individual collecting the data to appraise everything for themselves. This is the opportunity to see something unexpected or simply to notice that not all is as intended (indicating a flaw somewhere along the line). So, each individual experimenter must attain sufficient expertise and awareness of details and the bigger picture themselves in order to be well-placed to obtain reliable results and new insights.

A second mentor is important to address many complementary aspects of training, including all of the myriad aspects of technical execution of experiments, through direct demonstration and careful observation The second mentor hopefully also is self-confident enough to model another key characteristic- looking at results from a different perspective and arguing conclusions with the PI or the team.

Who were your most influential mentors/role models in science and what did you learn from them?

My PhD co-incided with early days of recombinant DNA cloning and, by chance, I was able to learn both the techniques and confidence (an expectation of mastery and further development) from cutting-edge leaders (Flavell, Grosveld) in a neighboring lab. My PhD mentor (Alan Smith) set this up and provided a good project with guidance, together with plenty of freedom to act independently over long periods of time. One of his favorite observations was that there is an optimum balance in most settings (for instance, in setting up any in vitro biochemical reaction)- too much can be as big a deficit as too little. The virtue of this advice is that it is very general- you just need to admit that it may apply to the question in hand. He also noted my preference for pursuit of a problem and thorough understanding in preference to writing papers at suitable intervals (but transmitting that information did not markedly alter the underlying behavioral trait). My post-doc project was not within the major thrust of the lab and so I was left (by Gerry Rubin) to operate extremely independently with the benefit of excellent support re lab organization and equipment, and a dozen contemporaries (mostly post-docs) learning from each other, mostly in terms of methodology. Distributed, incidental peer mentoring is pretty effective, in my opinion. I was definitely still learning about Drosophila genetics and developmental biology when starting as a PI here, and probably had the most helpful and productive interactions with Gary Struhl when our work converged at the PKA/Hh nexus- my first intensive experience of rigorous developmental genetics to produce a story.

Can you recommend courses/lectures in Columbia University that would be most beneficial for students/postdocs?

Our Biological Sciences PhD program has a required and closed Writing course that punches above its weight, and I suspect the same would be true of other courses on Writing or communication skills. Similarly, for any course that examines research papers of interest with extreme rigor (taking plenty of time). Regarding stem cells, I teach a course that is geared towards undergraduate seniors or Masters students; serving as a teaching assistant for that course would likely be very instructive.

What would be your career advice for students/postdocs?

For those on a road to academic research, they will already have the most essential ingredient of being really interested in progressively building greater understanding, and they are probably also aware that very many areas of research would be of interest if explored. My main advice would be to qualify yourself to be able to develop a career. This involves developing important research capabilities and making potential employers aware of those capabilities. Undoubtedly, that involves becoming exceptionally good in one, two or three approaches (“techniques”), completing a notable story or stories, and communicating in a way that makes clear you are the original source of your words and ideas, you think on your feet and can interact in those ways on numerous topics other than your own research. To do all that, you need to start with a suite of strong attributes, work hard, think hard, read fairly extensively and continually trial ideas privately and publicly. On top of that, I think that joining a lab with special technological expertise can be a great advantage.

Are you accepting rotating students at the moment?

Yes

Lab management

How do members of your lab celebrate accomplishments?

Probably, largely without my knowledge. Becoming a year older is often celebrated with cake or other life-shortening materials. Completing a PhD is of course celebrated as a notable achievement with a clear focal point of the thesis defense. The publication process takes most of the joy and celebratory opportunity out of scientific discovery and achievement because it is so long and almost inevitably involves a series of negative or intermediate decisions, with a final concession to publish long after discovery and original formulation of conclusions were fresh. The first moment of realization (for conclusions that turn out later to be correct) remains, independent of publication process, but that is a prize separate from public celebration.

What is the key to running a successful lab?

An enormous number of things, including implementing most from the very beginning because “success” is very difficult in the absence of continued momentum. I only practice a fraction. All of those elements are, of course, intended to make the most of the key raw ingredients – people, and in the hope of initiating a process where new lab members are attracted by those already present and how they interact to form a lab. If “successful” means highly productive, I think one ingredient is ambition to do just that, and that requires covering all the bases. If it primarily means continually making rigorous progress (even if on a smaller scale), or training others, or learning, or enjoying research (rather than all of these things with no compromises), the set of answers would be different. I think every one of these ambitions requires a huge amount of sustained time and attention; that may only be practical by prioritizing your natural preferences and abilities, rather than thinking about a generic formula or what you “should” do. In other words, you may have to concentrate on just a couple of laudable characteristics that you consider most important or achievable. Fortunately, you are not the only one running the lab, and others are likely to pick up complementary objectives.

What was the most exciting part about starting your new lab?

Having people join the lab and start to make progress. The prior period of relying primarily on your own hands for every scrap of new insight is inappropriately long, so the opportunity to think and act more broadly is long overdue.

Stem Cell Directions:

What are the most important recent developments in the stem cell field?

Since our work only relates to adult stem cells, I will answer only with respect to that. Aside from the obvious technological developments of CRISPR techniques, single cell sequencing and better imaging, I would say that the ability to follow events through live imaging is THE most powerful attribute that can be brought to bear on any paradigm. The limitations of indirect inferences from observations at fixed times are considerable, under-estimated and have, I believe, led to many mis-conceptions, some still to be sorted out.

Which stem cell conferences does your lab attend?

The most common is actually the annual Drosophila research conference- not a stem cell conference at all. While commonalities with a stem cell community are beneficial, our research fits more broadly within developmental biology, and in some cases, cell biology. More relevant insights and raw materials for our research has come from those broader fields and I believe it is beneficial to see stem cell research within the context of broader fields.

CSCI:

What was the main reason of you joining CSCI? What are the beneficial aspects of CSCI membership for your lab?

Communication connections with individuals outside my Department and with overlapping interests. Many years ago, I imagined helping to bring campuses together to some degree through CSCI-related efforts (training programs and courses, mainly). Physical separation is, however, a substantial barrier. Our lab has relatively few interactions with CSCI- mainly just the occasional seminar, plus “annual retreats.”

What do you plan to bring to the CSCI community?

I still have some hopes for contributing to educational and training material and activities. Intellectually (in collegial or advising interactions), I believe that simple paradigms, like ours, will always have something to offer, including a different perspective from the majority of CSCI research. In terms of scientific interactions I think the common ground is sufficient throughout CSCI that I, for example, could always ask relevant critical questions as part of a training exercise or in response to a talk.