SSSCA1 Human

What is SSSCA1 and what is its genomic location?

From a research perspective, it's important to note that SSSCA1 has alternative nomenclature in various databases:

What is the structural composition of SSSCA1 protein?

The crystal structure of SSSCA1's N-terminal domain has been determined at 2.3 Å resolution, revealing a zinc finger fold classified as a zinc ribbon domain type 2 (ZNRD2). Each protomer binds one zinc ion coordinated by four conserved cysteines (human residues Cys53, Cys56, Cys70, and Cys73) structured around four antiparallel β-sheets. Notably, residues 1-12 and 78-111 could not be modeled in the electron density map, corresponding to regions predicted to be intrinsically disordered .

The C-terminal domain serves a dual function: it acts as both the interaction site for Tankyrase 1 (TNKS1) binding and as a nuclear export signal. This functional duality represents an important consideration for experimental design when studying SSSCA1 localization and interaction networks .

How are anti-SSSCA1 antibodies detected in clinical samples?

Detection of anti-SSSCA1 antibodies in patient samples can be accomplished through immunoprecipitation of 35S-methionine-labeled protein generated by in vitro transcription and translation. This technique provides high specificity and sensitivity for detecting autoantibodies in serum samples from patients with autoimmune conditions .

For research laboratories, ELISA-based methods are also available for detecting human SSSCA1, with commercial kits offering detection ranges of 0.313-20 ng/ml and sensitivities below 0.188 ng/ml. These assays typically employ a sandwich ELISA format with colorimetric detection methods. When conducting research with such assays, optimal dilutions should be determined empirically for each specific application and sample type .

How does SSSCA1 interact with Tankyrase 1 (TNKS1)?

SSSCA1 directly interacts with Tankyrase 1 (TNKS1) through its C-terminal domain. This interaction has been experimentally verified and mapped to specific regions of both proteins. The C-terminal H2 helix region of SSSCA1 is critical for binding to TNKS1, as demonstrated by mutation studies where changing leucine residues to alanine (SSSCA1-H2 Leu/Ala) abolished this interaction .

From the TNKS1 perspective, the binding site for SSSCA1 has been mapped to the ankyrin repeat cluster 2 (ARC2) within the ankyrin repeat domain (ARD). This was determined through deletion mapping experiments where TNKS1 constructs lacking the sterile alpha motif (SAM) and poly-ADP-ribose polymerase (PARP) domains still maintained SSSCA1 binding, while deletion of the ARD abolished the interaction .

When designing experiments to study this interaction, researchers should consider:

• Using co-immunoprecipitation with either tagged SSSCA1 or TNKS1

• Employing domain-specific mutants to validate interaction sites

• Considering the impact of subcellular localization on interaction dynamics

What experimental approaches are most effective for studying SSSCA1 subcellular localization?

To effectively study SSSCA1 subcellular localization, researchers should employ a combination of techniques:

• Fluorescent protein tagging: The human gene of SSSCA1 can be cloned in frame with an 8x His-GFP tag and transiently transfected into cancer cell lines. Following 18 hours of expression, subcellular localization can be observed through confocal microscopy .

• Nuclear export signal (NES) validation: Since SSSCA1 contains a functional nuclear export signal in its C-terminal domain, researchers should consider using leptomycin B (an inhibitor of nuclear export) to confirm NES functionality. Combined with mutational analysis of the NES sequence, this approach can elucidate the regulation of SSSCA1 nuclear-cytoplasmic shuttling .

• Co-localization studies: Given SSSCA1's reported association with centromeres during mitosis, co-immunostaining with centromere markers during different cell cycle phases can provide insights into its dynamic localization patterns .

What is the association between anti-SSSCA1 antibodies and cancer in systemic sclerosis (SSc) patients?

Recent research has identified a significant association between anti-SSSCA1 antibodies and cancer in systemic sclerosis patients. In a case-control study of 414 patients (209 with SSc and cancer, 205 with SSc without cancer), 31 patients (7%) were anti-SSSCA1 antibody positive. The data revealed several important findings:

• Patients with cancer were significantly more likely to be anti-SSSCA1 positive compared to those without cancer (11% vs. 4%, P = 0.018) .

• After adjustment for potential confounders, patients with anti-SSSCA1 antibodies demonstrated an increased risk of cancer (odds ratio 2.46, 95% CI 1.06-5.70) compared to anti-SSSCA1-negative patients .

• Among patients with cancer, there was a trend toward longer cancer-SSc interval in anti-SSSCA1-positive patients compared to anti-SSSCA1-negative patients .

This evidence suggests that anti-SSSCA1 antibody status may serve as a potential cancer biomarker in SSc patients and should be considered in clinical research protocols examining the cancer-autoimmunity interface.

What clinical manifestations are associated with anti-SSSCA1 antibody positivity in SSc patients?

Anti-SSSCA1 antibody-positive SSc patients display distinct clinical manifestations compared to antibody-negative patients. Research has identified several key clinical features:

• Cardiovascular involvement: Anti-SSSCA1-positive patients demonstrate a lower minimum ejection fraction and a trend toward more severe heart involvement .

• Pulmonary function: These patients typically present with a lower baseline diffusing capacity of the lungs for carbon monoxide (DLCO) percent predicted .

• Peripheral vascular disease: Anti-SSSCA1-positive patients are more likely to have severe Raynaud's phenomenon .

When designing clinical studies involving SSc patients, researchers should consider stratifying by anti-SSSCA1 antibody status and specifically assess these clinical domains, as they may represent a distinct clinical phenotype with implications for prognosis and treatment response.

What are the optimal conditions for expressing and purifying recombinant human SSSCA1?

When expressing and purifying recombinant human SSSCA1 for structural and functional studies, researchers should consider the following methodological approaches:

• Expression systems: For structural studies, the N-terminal domain (ZNRD2, residues 1-77) has been successfully expressed in E. coli BL21(DE3) cells. For full-length protein or functional studies involving post-translational modifications, mammalian expression systems may be preferable .

• Purification strategy: His-tagged purification approaches have been successful, with affinity chromatography followed by size exclusion chromatography yielding protein suitable for crystallographic studies .

• Buffer considerations: Given the zinc-binding properties of the N-terminal domain, buffers should be optimized to maintain zinc coordination, typically including reducing agents to prevent cysteine oxidation.

• Protein stability: SSSCA1 demonstrates regions of intrinsic disorder, which may impact long-term stability. Storage conditions should be empirically determined, with flash-freezing in the presence of glycerol or similar cryoprotectants often proving effective.

How can protein-protein interactions of SSSCA1 be systematically mapped?

To comprehensively map the protein-protein interactions of SSSCA1, multiple complementary approaches should be employed:

• Affinity purification-mass spectrometry (AP-MS): This approach has been successfully used to identify TNKS1 as a direct binding partner of SSSCA1. Expressing tagged SSSCA1 in relevant cell lines followed by pull-down and mass spectrometric analysis can identify novel interaction partners .

• Yeast two-hybrid screening: This can serve as a complementary approach to AP-MS, potentially identifying interactions that may be transient or context-dependent.

• Proximity-based labeling: BioID or APEX2-based approaches, where SSSCA1 is fused to a proximity-based labeling enzyme, can identify proteins in the vicinity of SSSCA1 in living cells, providing spatial context to interaction networks.

• Domain-specific interaction mapping: As demonstrated with the C-terminal domain's interaction with TNKS1, domain-specific constructs should be used to map the specific regions mediating individual protein-protein interactions .

How might SSSCA1's role in the Wnt signaling pathway be further elucidated?

SSSCA1 has been implicated in the Wnt signaling pathway through mass spectrometric and proteomic studies. To further elucidate its role, researchers should consider:

• Pathway perturbation analysis: Using CRISPR-Cas9 knockout or knockdown approaches to modulate SSSCA1 expression followed by assessment of Wnt pathway activity using reporter assays (e.g., TOPFlash assays).

• Interaction with canonical Wnt pathway components: Given SSSCA1's interaction with TNKS1, which is known to regulate axin degradation in the Wnt pathway, specific investigations into how SSSCA1 might modulate TNKS1's effects on axin stability would be valuable.

• Context-dependent functions: Examining SSSCA1's role in the Wnt pathway across different cell types and disease states, particularly in cancer models where Wnt signaling is frequently dysregulated.

What are the implications of SSSCA1's crystal structure for drug discovery efforts?

The determined crystal structure of SSSCA1's zinc ribbon domain at 2.3 Å resolution provides valuable insights for structure-based drug design approaches:

• Zinc coordination sites: The four conserved cysteines (Cys53, Cys56, Cys70, and Cys73) that coordinate zinc represent potential targets for metal-binding inhibitors or compounds that could disrupt domain stability.

• Protein-protein interaction interfaces: With the identification of the C-terminal domain as the TNKS1 binding site, structure-guided design of peptide mimetics or small molecules that could disrupt this interaction becomes feasible.

• Nuclear export signal targeting: The dual function of the C-terminal domain as both a TNKS1 binding site and nuclear export signal suggests that compounds targeting this region might modulate both SSSCA1's subcellular localization and its interaction network.

When designing such studies, researchers should consider the intrinsically disordered regions (residues 1-12 and 78-111) that could not be resolved in the crystal structure, as these might adopt ordered conformations upon binding to partners or ligands .