SSPsyGene Consortium – Frequently Asked Questions (FAQs)

Answers to frequently asked questions about the following two funding opportunity announcements (FOAs): Scalable and Systematic Neurobiology of Psychiatric and Neurodevelopmental Disorder Risk Genes: Assay and Data Generation Centers (RFA-MH-22-111) and Scalable and Systematic Neurobiology of Psychiatric and Neurodevelopmental Disorder Risk Genes: Data Resource and Administrative Coordination Center (RFA-MH-22-110).

• Gene Selection and Numbers
• Choice of Experimental Systems/Species
• Choice of Assays
• Overarching Topics

Gene Selection and Numbers

Question: What are the target number of genes to be investigated within the SSPsyGene Consortium over the 5-year project period?
Response: It is anticipated that each Assay and Data Generation Center (ADGC) supported through the SSPsyGene Consortium will characterize the full set of 100-250 genes selected by the Consortium in each proposed assay over the course of the 5-year project period. It is understood that some targeting approaches may not be appropriate for all genes or all assays (e.g., a constitutive homozygous null for one gene may be lethal in a model organism), but the expectation is that, to the extent feasible, the full gene set will be interrogated in each assay.

Question: Is the list of genes available? If not, how will the genes to be assessed by the SSPsyGene Consortium be selected? Does each applicant need to come up with a separate plan for gene list selection?
Response: While the long-term goal of the SSPsyGene Consortium is to build a comprehensively annotated resource describing the CNS function of all neurodevelopmental and psychiatric disorder (NPD) risk genes, the target for the initial phase of the initiative is 100-250 protein coding genes. The Data Resource and Administrative Coordination Center (DRACC) will be responsible for convening the members of the Consortium, within 3 months of award, to prioritize a set of ~100-250 genes associated with NPDs for assessment within the SSPsyGene Consortium. Several criteria should be taken into account when prioritizing genes:

• The exact number of genes selected will depend on feasibility given costs, potential assays, and the available timeframe.
• Initial characterization efforts will focus primarily on null alleles. To maximize the potential disease relevance of null effects, genes with a genome-wide significant excess burden of loss-of-function mutations in NPDs such as intellectual disability/developmental delay, autism spectrum disorder, and schizophrenia will be the focus. The Consortium will also determine the need to assess heterozygous and homozygous states, and develop a rigorous, data-informed approach for selecting patient variants to pilot assays interrogating an allelic series from a small, select subset of NPD risk genes. The DRACC will be responsible for leading the gene prioritization process, including engaging external scientific consultants (e.g., with statistical genetics expertise) to serve with key investigators on the SSPsyGene Consortium Coordinating Committee (CCC).
• Additional factors to consider are expression within the experimental systems, conservation across species, developmental specificity, balanced representation across disorders, existing knowledge of gene function (with less studied genes to be prioritized), and alignment with the recommendations from the Report of the National Advisory Mental Health Council Workgroup on Genomics.

As noted in the application instructions, applicants for both the DRACC and the ADGCs should describe a proposed approach for selecting a set of 100-250 NPD risk genes, with the understanding that strategies proposed by the DRACC and ADGCs will be discussed by the DRACC and the CCC in finalizing a gene list after award.

Question: Can I do unbiased genome-wide screening? Can I include analysis of copy number variants (CNVs)?
Response: The goal of this initiative is to fill a very specific knowledge gap area, which is that many genes are now associated with NPD risk, but their basic neurobiology remains poorly understood. SSPsyGene is designed to provide a collaborative and efficient framework for identifying biological function beyond current small-scale and often ad hoc single gene efforts in order to generate a standardized, experimentally derived, functional catalog of NPD risk genes. This basic neurobiology resource would provide a fertile foundation for future studies into NPD disease mechanisms. Because of this, broader genome-wide screens and analysis of larger multigenic intervals (CNVs) associated with disease risk are beyond the scope of this initiative.

Question: Are null alleles the focus for the initiative? Many of these genes may be essential genes, making analysis of null alleles difficult in some contexts. What about heterozygous versus homozygous null?
Response: Initial characterization will focus primarily on null alleles. To maximize the potential disease relevance of null effects, genes with a genome-wide significant excess burden of loss-of-function mutations in NPDs such as intellectual disability/developmental delay, autism spectrum disorder, and schizophrenia will be the focus. The Consortium will also determine the need to assess heterozygous versus homozygous states, and develop a rigorous, data-informed approach for selecting patient variants to pilot assays against an allelic series from a small, select subset of NPD risk genes.

It is understood that some targeting approaches may not be appropriate for all genes or all assays (e.g., a constitutive homozygous null for one gene may be lethal in a model organism). It is also understood that some targeting approaches are more costly, time- and labor-intensive than others. As an initial first pass into gene function, applicants can propose highly scalable genetic perturbation approaches that do not involve individual stable mutants (e.g., CRISPR interference).

As noted in the Application Instructions for the ADGCs, applicants are required to describe how alleles will be produced along with the rationale for this choice. Applications may propose to generate new null alleles, or may use existing resources (e.g., available libraries of CRISPR/Cas9 alleles in iPSCs that can be differentiated into systems for assays; knock-out animal resources such as the Knockout Mouse Project, Zebrafish International Resource Center). Proposed approaches need not be strictly limited to actual genetic knockouts, as long as the alleles can be functionally equivalent to a null (i.e., do not produce functional protein) and otherwise aid in attaining the FOA goals. Applications should describe how alleles will be ascertained/QC'd to ensure they are null and stable enough to work in assays and how they will assess dosage sensitivity. Details for generating and validating an allelic series of patient variants from a small subset of genes should also be provided, as well as a discussion of potential artifacts or confounders (e.g., genetic compensation for certain alleles; cases in which the homozygous null is cell lethal in their assays; cases where heterozygous null effects are not detected in selected assays, degree of cross-species conservation) and how they will be addressed.

Choice of Experimental Systems/Species

Question: Is any particular experimental system required as a component of an ADGC application? Or is the RFA agnostic to the model organism/experimental system? I noticed that mouse is missing from the model organisms of interest. Are proposals focused on assays in rodent systems a “no go”?
Response: The intention of the SSPsyGene initiative is to leave the types of assays and models open to encourage a diversity of approaches and technologies. Applications are only constrained by the primary focus on molecular and cellular levels of biological organization, and the throughput capabilities of the proposed system to rigorously and systematically characterize 100-250 genes within the possible budget. Different experimental systems may be more or less amenable to a specific functional/phenotypic readout. It is possible that applications proposing to utilize pluripotent cell-based paradigms or model organisms with a short generation time will be most feasible within such budget and throughput constraints. However, applicants are welcome to propose projects using other model systems and encouraged to carefully justify project feasibility in such applications.

Question: Will each ADGC be expected to generate the null alleles within their model system?
Response: The SSPsyGene Consortium is expected to be collaborative, including the sharing of resources and reagents, when feasible. However, ultimately each ADGC will be responsible for targeting the common set of genes in their chosen system(s), taking advantage of existing model organism and cellular resources where available.

Choice of Assays

Question: Should an applicant discuss the inherent limitation of proposed assays for interrogating all of the genes proposed by the SSPsyGene Consortium? Each assay will have limitations that may make it technically challenging and, in some cases, impossible to interrogate every gene and it will not be possible to know all of the potential issues in advance, as the gene list has not yet been developed?
Response: It is understood that some genes may not be appropriate for all assays (e.g., null allele cannot successfully be generated in a particular experimental system, gene is not expressed in a particular cell type) and that it will not be possible to know all of the potential issues until the Consortium has selected the genes to be interrogated. As with any application, it is important to consider potential pitfalls and alternatives in the proposed experimental plan. For each assay, applicants are encouraged to discuss technical limitations of the approach that may limit success for certain gene targets and discuss potential alternatives approaches that could be employed.

Question: If an investigator has already created and characterized a large number of null alleles of relevance to neuropsychiatric disorders, would it be reasonable for a proposal to include these assays to complete the data set? Or are you looking only for unique datasets that have not been done or published to date?
Response: It would be reasonable to include already-generated null alleles and associated assays in an ADGC proposal to advance systematic characterization of the risk genes for that functional measure. Existing data using the assay may be viewed as preliminary data to support feasibility of conducting the assay at scale and support the significance of the functional measure for characterizing gene function. Incorporation of novel assays that have not yet been applied broadly to assess the function of NPD risk genes is also encouraged.

Question: Will the FOA support characterization of gene function across various neuronal and non-neuronal cell types or are experimental systems/assays limited to neurons?
Response: Assays to assess gene function in any CNS cell type are allowable under the SSPsyGene FOAs.

Question: Is the focus on molecular/cellular levels of analysis? Are assays targeting neuronal communication or neural circuit function within the scope of the FOA?
Response: The initial focus for the SSPsyGene Consortium will be on the implementation of high-throughput molecular and cellular assays. Ultimately, the goal of the initiative is to broadly characterize genes across a variety of fundamental CNS functions including developmental processes, cell morphology, neurite motility, intrinsic membrane properties, synaptic physiology, inter- and intra-cellular signaling, circuit dynamics, natural behaviors, or other neural phenotypes. While the primary focus at this time is at the molecular/cellular scale, assays may include CNS-relevant phenotypes across scales of biological organization from molecular and cellular to physiological and organismal, if they are amenable for scalable and systematic investigations.

Question: Should applications propose initial validation of results from the high throughput assays? Do these need to be done at the same scale/throughput?
Response: While highly scalable measures to support systematic characterization of gene function are the primary focus of the initiative, applications should incorporate plans for validating key results to ensure rigor. While maintaining scalability/throughput throughout the validation process is encouraged, applicants may propose lower throughput approaches at later stages of validation with appropriate justification as to the need and selective use.

Question: Is it preferred that applications focus on a single level of analysis or should applicants propose assays across multiple levels? Can one propose a single assay across multiple model organisms?
Response: Each application must propose projects that address at least one of the high priority assay areas listed described in the ADGC FOA (Molecular Assays; Cellular, Circuit, or Systems/Organismal Assays). Applicants have the flexibility to select any assay category or may opt to propose assays in multiple areas or across multiple experimental systems / species. Assays should be considered based on their throughput and the utility of information they provide to advance understanding of the neurobiological function of NPD risk genes. Proposals to carry out multiple complementary assays within each priority area or across areas are of interest.

Question: Are behavioral assays of interest? How about behavioral assays in conjunction with a priority assay listed in the RFA?
Response: The primary focus of the SSPsyGene Consortium is on high-throughput molecular and cellular analyses. While robust, scalable and reproducible behavioral phenotyping may be proposed, successful RM1 applications will likely also have a strong focus on molecular and/or cellular assays. Further, with the goal of achieving a systematic characterization at scale, each of the RM1 awardees will be expected to complete rigorous functional characterization all of the 100-250 genes that the Consortium appoints to the common core gene set within the project period. Applications proposing to utilize pluripotent cell-based paradigms or model organisms with a short generation time may be most feasible within such budget and throughput constraints.

Overarching Topics

Question: How do we describe interactions between the ADGC and DRACC when we don’t know who the partners will be?
Response: For the ADGC (RM1 applications), applicants should describe their internal laboratory information management system and data/metadata processing pipeline such that it will allow systematic control of data flow internally, along with accurate and efficient transfer of such information to another party (e.g., via the DRACC). Applicants should also discuss past experiences (if any) in working as part of interdisciplinary teams and/or large collaborative research efforts, as well as plans for maintaining a high level of Consortium engagement in strategic planning, including involvement with the DRACC. More detail is found in Section IV, Research Strategy, part “3) Data Management and Analysis” and “4) Consortium Involvement and Integration”. Applications that do not include this will be considered non-responsive.

Question: What is the anticipated or ideal deliverable for the SSPsyGene Consortium?
Response: The SSPsyGene Consortium aims to produce a systematic annotation of basic neurobiological function of genes associated with risk for NPDs. The resulting phenotypic data will be integrated across modalities, levels of organization, and genes to create a harmonized, integrated knowledge base needed to substantively advance our understanding of basic gene function and, ultimately, make robust inferences into potential shared and unique disease mechanisms.