Recombinant Mouse Protein FAM204A (Fam204a)

What is Mouse Protein FAM204A and what genomic information is available? [BASIC]

Mouse FAM204A (Family with sequence similarity 204 member A) is encoded by the Fam204a gene, also known by synonyms 2310065H12Rik, 2610015K05Rik, and D19Ertd737e. The gene has multiple transcript variants documented in RefSeq (NM_029648, NM_001358271, NM_001358272, NM_001358273, NR_152390, NR_152391). The protein has a UniProt ID of Q8C6C7 and a Locus ID of 76539 .

FAM204A belongs to a protein family characterized by the InterPro domain IPR037690. The zebrafish ortholog (fam204a) has been classified as a protein-coding gene located on chromosome 13, further supporting evolutionary conservation of this gene family .

What is currently known about the domain structure and protein characteristics of FAM204A? [BASIC]

Based on available data, mouse FAM204A contains the InterPro domain IPR037690 (Protein FAM204A family). The zebrafish ortholog indicates a protein length of 234 amino acids . While direct structural information about mouse FAM204A is limited in current literature, analysis of related FAM proteins suggests potential features.

How is FAM204A expressed during development or in specific tissues? [ADVANCED]

Current literature provides limited direct evidence about FAM204A tissue-specific expression patterns. For researchers investigating expression patterns, recommended approaches include:

• RT-qPCR analysis across multiple tissue types and developmental stages

• Western blot analysis using validated antibodies

• RNA-seq data mining from public databases

• In situ hybridization for spatial expression analysis

Related FAM proteins show tissue-specific expression patterns – for example, FAM208A (D14Abb1e) shows strong expression in skin, brain, and testis , while FAM20 family members show differential expression during hematopoietic differentiation . Similar methodologies could be applied to characterize FAM204A expression.

What are recommended approaches for detecting endogenous FAM204A in mouse tissues? [BASIC]

For researchers aiming to detect endogenous FAM204A, a multi-method approach is recommended:

When selecting antibodies, researchers should verify specificity through appropriate controls such as knockdown validation, as demonstrated with FAM208A detection where siRNA treatment confirmed antibody specificity .

What genetic modification strategies are available for studying FAM204A function? [ADVANCED]

Current tools for genetic manipulation of Fam204a include:

• CRISPR activation (CRISPRa) technology for upregulating endogenous Fam204a expression. Commercial kits include three gRNA vectors targeting Fam204a, a CRISPRa-Enhancer vector, and appropriate controls .

• CRISPR knockout systems employing non-homology mediated gene editing, enabling loss-of-function studies .

When designing genetic manipulation experiments, researchers should consider:

• Cell-type specific effects of FAM204A modulation

• Potential compensatory mechanisms by related genes

• Phenotypic analysis at multiple levels (molecular, cellular, physiological)

• Appropriate control conditions (scrambled gRNAs, empty vectors)

The efficacy of CRISPR-based activation can be affected by factors including "nucleosome occupancy status, chromatin structure and gene expression level of the target" , requiring optimization for specific experimental systems.

What recombinant expression strategies are optimal for producing functional mouse FAM204A protein? [ADVANCED]

While the literature doesn't provide specific optimization parameters for FAM204A expression, researchers can apply established approaches based on predicted protein characteristics:

Expression system selection should be guided by:

• Predicted post-translational modifications

• Experimental application requirements

• Scale of protein production needed

• Budget and time constraints

What approaches can elucidate potential functions of FAM204A in cellular processes? [BASIC]

To determine FAM204A function, researchers could employ these methodological approaches:

• Subcellular localization studies using fluorescent protein fusions or immunolocalization

• Interactome analysis using techniques such as:

  • Affinity purification followed by mass spectrometry

  • Yeast two-hybrid screening

  • Proximity labeling (BioID, APEX)

• Phenotypic analysis following genetic perturbation:

  • Proliferation and cell cycle analysis

  • Differentiation capacity assessment

  • Transcriptomic changes via RNA-seq

For context, related FAM protein studies have yielded functional insights - FAM208A interacts with H3K9me3 (a chromatin mark associated with gene silencing) , and FAM20 family members play roles in regulating differentiation and function of hematopoietic tissues .

How can researchers investigate potential developmental roles of FAM204A? [ADVANCED]

Developmental biology approaches to study FAM204A might include:

• Temporal expression analysis during embryonic development

• Conditional knockout models using tissue-specific or temporally regulated Cre lines

• Chimeric embryo analysis using FAM204A-deficient ES cells

• In vitro differentiation models using embryonic stem cells or induced pluripotent stem cells

When interpreting developmental phenotypes, lessons from related FAM proteins are instructive. For example, homozygous mutations in D14Abb1e (FAM208A) resulted in embryonic lethality beyond gastrulation, demonstrating its essential role in development . Analysis of heterozygous animals revealed effects on transgene expression at the RNA level, suggesting dosage sensitivity .

What experimental approaches can help determine if FAM204A has potential roles in gene regulation? [ADVANCED]

To investigate potential gene regulatory functions, researchers might employ:

• Chromatin immunoprecipitation followed by sequencing (ChIP-seq) to identify genomic binding sites

• RNA-seq following FAM204A perturbation to identify regulated genes

• ATAC-seq to assess chromatin accessibility changes

• Protein-protein interaction studies focusing on known transcriptional regulators

These approaches should be complemented with mechanistic validation experiments such as reporter assays or directed mutagenesis of functional domains.

For comparison, FAM208A (D14Abb1e) was found to interact with H3K9me3, a chromatin mark associated with gene silencing , while FAM20A expression varied during hematopoietic differentiation, suggesting potential regulatory roles in this process .

How does mouse FAM204A compare to human FAM204A and orthologs in other species? [BASIC]

Comparative analysis of FAM204A across species provides evolutionary context:

• Zebrafish fam204a encodes a protein of 234 amino acids, classified in the Protein FAM204A family (IPR037690)

• Evolutionary analysis of other FAM protein families shows patterns of gene duplication and diversification

• FAM20 family demonstrates expansion from a single gene in simpler organisms (C. elegans) to multiple genes in vertebrates

When conducting comparative studies, researchers should:

• Use multiple sequence alignments to identify conserved domains

• Consider syntenic relationships when confirming true orthologs

• Examine expression patterns across species to identify conserved regulatory mechanisms

What insights can be gained from studying evolutionary relationships between FAM204A and other FAM proteins? [ADVANCED]

Evolutionary analysis of FAM protein families can provide functional insights:

• Phylogenetic tree construction to determine relatedness to functionally characterized FAM proteins

• Identification of conserved sequence motifs that may indicate shared functional domains

• Analysis of selection pressure (dN/dS ratios) across different domains

• Correlation of gene duplication events with evolutionary innovations

The expansion observed in the FAM20 family, which evolved through gene duplication events in both vertebrates and invertebrates , suggests similar analyses could provide insights into FAM204A evolution and potential functional specialization.

What purification strategies are most effective for recombinant mouse FAM204A? [BASIC]

Optimal purification strategies for recombinant FAM204A should consider:

A typical purification workflow might include:

• Expression screening in selected system

• Initial affinity purification using tagged construct

• Tag removal (if necessary) followed by secondary purification

• Quality control analysis (see section 5.2)

What quality control measures should be applied to purified recombinant FAM204A? [ADVANCED]

Comprehensive quality control for recombinant FAM204A should include:

• Purity assessment:

  • SDS-PAGE with Coomassie or silver staining

  • Size exclusion chromatography

  • Mass spectrometry

• Identity confirmation:

  • Western blotting with FAM204A-specific antibodies

  • Peptide mass fingerprinting

  • N-terminal sequencing

• Structural integrity:

  • Circular dichroism to assess secondary structure

  • Thermal shift assays to evaluate stability

  • Dynamic light scattering to detect aggregation

• Functional validation:

  • Binding assays with known interaction partners (once identified)

  • Activity assays based on discovered functions

  • Comparison with native protein where possible

Researchers should establish acceptance criteria for each quality parameter based on the intended experimental application of the recombinant protein.