Recombinant Human C-C motif chemokine 7 protein (CCL7) (Active)

What is the molecular structure of human CCL7 protein?

Human CCL7 consists of 99 amino acids, which includes a 23-amino acid signal peptide. The mature and active form contains approximately 76 amino acids after cleavage of the signal peptide . Unlike most chemokines that typically form dimers in concentrated solutions, CCL7 predominantly exists in a monomeric form, which represents a unique structural characteristic . The protein belongs to the C-C subfamily of chemokines, characterized by having two adjacent cysteine residues that are critical for its tertiary structure .

What are the common synonyms and identifiers for CCL7?

CCL7 has been known by several different names throughout scientific literature, which can sometimes create confusion when searching databases. The protein is also referred to as:

• MCP3 or MCP-3 (Monocyte-Chemotactic Protein 3)

• MARC

• FIC

• NC28

• SCYA6

• SCYA7

The gene encoding CCL7 has been assigned the official locus symbol SCYA7 in the human genome .

Where is the CCL7 gene located in the human genome?

The CCL7 gene is located on chromosome 17q11.2-q12, forming part of a cluster with other chemokine genes, particularly those encoding the MCP subset of CC chemokines . The genomic structure consists of three exons and two introns. The first exon contains a 5′-untranslated region (5′-UTR), the information for the signal sequence (23 amino acids), and the mature protein's first two amino acids. The second exon encodes amino acids 3–42 of the mature protein, while the third exon encodes the C-terminal region and contains a 3′-UTR with destabilizing AU-rich sequences and a polyadenylation signal .

What post-translational modifications occur in CCL7?

CCL7 can exist in four different glycotypes with molecular weights of 11, 13, 17, and 18 kDa when expressed in COS cells . These glycosylation patterns may affect protein stability, receptor binding affinity, and biological activity. Understanding these modifications is crucial for researchers working with recombinant CCL7, as expression systems may yield proteins with different glycosylation patterns compared to naturally occurring CCL7.

Which receptors does CCL7 interact with and what are their distributions?

CCL7 mediates its biological effects through binding to multiple C-C chemokine receptors, including CCR1, CCR2, CCR3, CCR5, and CCR10 . These receptors belong to the G protein-coupled seven-transmembrane receptor family. This broad receptor binding profile makes CCL7 unique among chemokines and explains its diverse biological activities. Additionally, CCL7 can interact with cell surface glycosaminoglycans (GAGs) present on virtually all animal cell surfaces, which may facilitate the formation of chemotactic gradients necessary for directed cell migration .

How does receptor binding specificity differ between CCL7 and related chemokines?

While CCL7 shares structural similarities with other CC chemokines, particularly CCL2 (formerly MCP1), its receptor binding profile is broader . This expanded receptor interaction capability allows CCL7 to attract a wider range of leukocytes compared to more selective chemokines. When designing experiments to study specific chemokine functions, researchers should carefully consider this overlap in receptor usage, as it may complicate the interpretation of results, especially in systems where multiple chemokines are present.

What intracellular signaling pathways are activated by CCL7?

CCL7 binding to its receptors initiates signaling through G protein-coupled pathways, similar to other chemokines. Although the search results don't provide specific details about CCL7 signaling, related chemokine receptors like CCR7 activate ERK1/2 and other signaling molecules . The temporal profile of these signaling events can vary significantly between different chemokines, even when they bind the same receptor. For instance, some chemokines induce prolonged signaling while others produce more transient responses .

What are the recommended methods for studying CCL7-induced cell migration?

When designing chemotaxis assays with CCL7, researchers should consider:

• Gradient steepness: Different cell types respond differently to steep versus shallow chemokine gradients. Based on studies with related chemokines, the concentration gradient significantly impacts migration efficiency .

• 3D versus 2D migration: Three-dimensional chemotaxis assays often provide more physiologically relevant data than traditional two-dimensional transwell assays. In studies with dendritic cells and CCL19/CCL21, 3D assays revealed functional differences not apparent in 2D systems .

• Opposing gradients: When investigating preferential migration toward competing chemokines, researchers should establish opposing gradients. This approach has revealed important functional differences between related chemokines in immune cell migration .

• Chemotactic index measurement: Calculate the ratio of cells moving toward versus away from the chemokine source, rather than just measuring absolute migration, to obtain more meaningful quantitative data .

How should recombinant CCL7 be handled and stored to maintain activity?

While specific handling instructions for recombinant CCL7 are not directly provided in the search results, general principles for chemokine handling apply:

• Reconstitution: Recombinant chemokines should typically be reconstituted in sterile, buffered solutions with a carrier protein (such as 0.1% BSA) to prevent adhesion to tubes and loss of activity.

• Storage: Aliquot and store at -80°C to avoid repeated freeze-thaw cycles which can degrade protein activity.

• Working solutions: Prepare fresh working solutions for each experiment when possible, particularly for chemotaxis assays where chemokine activity is critical.

• Validation: Confirm biological activity through established functional assays, such as calcium mobilization or chemotaxis of responsive cell types, before using in complex experimental systems.

What is the role of CCL7 in inflammatory diseases?

CCL7 has been implicated in various inflammatory conditions, particularly Crohn's disease. Genome-wide association studies have identified CCL7 as a potential susceptibility locus for Crohn's disease, particularly in Ashkenazi Jewish populations . As a potent chemoattractant for macrophages and other inflammatory cells, CCL7 likely contributes to the pathogenesis of chronic inflammatory disorders through the recruitment of leukocytes to inflamed tissues .

How is CCL7 involved in cancer progression?

While the search results don't provide specific information about CCL7 in cancer, data on related chemokines suggest potential roles in tumor progression. Chemokines can be secreted by tumor cells to form an inflammatory environment conducive to tumor development . They may affect tumor proliferation, invasion, migration, and angiogenesis through autocrine and paracrine mechanisms involving multiple signaling pathways . These insights may guide research into the potential roles of CCL7 in cancer biology.

What is known about CCL7 in viral infections?

• Temporal expression patterns during infection

• Cell types producing CCL7 in response to viral stimuli

• Impact of viral proteins on CCL7 expression and function

• Potential targeting of CCL7 or its receptors by viral immune evasion mechanisms

What are the key methodological challenges when studying differential signaling through CCL7 receptors?

Investigating CCL7 receptor signaling presents several challenges:

• Receptor redundancy: CCL7 binds multiple receptors (CCR1, CCR2, CCR3, CCR5, CCR10), making it difficult to attribute signaling events to specific receptor interactions .

• Biased signaling: Different ligands for the same receptor may induce distinct signaling patterns (biased agonism). Studies with CCR7 ligands demonstrated that structurally related chemokines can induce temporally distinct patterns of ERK1/2 activation .

• Cell surface modifications: Sialic acid residues and other glycosylations can significantly impact chemokine-receptor interactions. Studies with CCR7 showed that neuraminidase treatment impaired ERK1/2 activation by some ligands but not others .

• Receptor mutations: Site-directed mutagenesis of key residues in chemokine receptors has revealed differential effects on ligand binding and signaling. For instance, mutations in the main binding pocket of CCR7 differentially affected responses to different ligands .

To address these challenges, researchers should consider experimental approaches that:

• Use receptor-specific antagonists or receptor knockout/knockdown models

• Employ multiple readouts of receptor activation to detect biased signaling

• Study effects of post-translational modifications on receptor function

• Use structure-guided mutagenesis to identify residues critical for ligand selectivity

How do glycosaminoglycan interactions affect CCL7 function in different experimental systems?

CCL7, like other chemokines, interacts with cell surface glycosaminoglycans (GAGs) . These interactions can significantly impact experimental outcomes:

• Gradient formation: GAGs immobilize chemokines on cell surfaces and extracellular matrix, creating stable gradients necessary for directional migration. In experimental systems lacking appropriate GAGs, chemotaxis may be impaired.

• Receptor access: GAG binding may alter chemokine conformations, affecting receptor recognition. Studies with tailless-CCL21 versus full-length CCL21 showed that GAG interactions can impair chemotaxis while enhancing certain signaling events .

• Protection from proteolysis: GAG binding can protect chemokines from proteolytic degradation, extending their half-life in experimental systems.

When designing CCL7 experiments, researchers should consider:

• The GAG composition of their experimental system

• Potential differences between recombinant and native CCL7 in GAG interactions

• The impact of soluble GAGs or GAG-binding inhibitors on CCL7 function

• Differences in GAG expression between cell types and tissues

What are the recommended approaches for studying CCL7 specificity in systems with multiple chemokines and receptors?

To dissect the specific contributions of CCL7 in complex systems:

• Receptor antagonists: Use specific receptor antagonists to block individual receptors in the CCL7 signaling network. Based on studies with other chemokine receptors, comparing the effects of different antagonists can reveal receptor-specific contributions to observed phenotypes .

• Gene editing approaches: CRISPR/Cas9 or RNAi technologies targeting CCL7 or its receptors can provide clean systems for studying specificity. Consider compensatory upregulation of related chemokines when interpreting results.

• Neutralizing antibodies: Specific antibodies against CCL7 can be used to selectively block its function without affecting related chemokines. Antibodies targeting specific epitopes have been used successfully to distinguish between full-length and truncated forms of chemokines .

• Chimeric receptors and ligands: Creating chimeras between CCL7 and related chemokines, or between its receptors and other chemokine receptors, can help identify domains responsible for specificity.

What are the critical considerations when analyzing CCL7 expression data in disease contexts?

When evaluating CCL7 expression in disease:

• Cell-specific expression: CCL7 is expressed by multiple cell types, including stromal cells, keratinocytes, airway smooth muscle cells, parenchymal cells, fibroblasts, leukocytes, and tumor cells . Bulk tissue analysis may mask important cell-specific expression changes.

• Post-translational regulation: CCL7 activity is regulated by proteolytic processing. It is an in vivo substrate of matrix metalloproteinase 2, which degrades components of the extracellular matrix . Measuring mRNA or even protein levels may not accurately reflect active chemokine levels.

• Receptor expression: The biological impact of CCL7 depends not only on its expression but also on the expression of its receptors on target cells. Comprehensive analysis should include receptor expression data.

• Functional redundancy: Other chemokines, particularly CCL2, share functional properties with CCL7 . Decreased CCL7 expression may be compensated by increased expression of functionally related chemokines.