CTSZ Human

What is CTSZ and what is its basic function in human biology?

Cathepsin Z (CTSZ), also named cathepsin P or cathepsin X, belongs to the cathepsin family comprising 11 members in humans. CTSZ contains an Arg-Gly-Asp motif responsible for binding to cell surfaces, mediating cell adhesion and migration after activation . This protein plays significant roles in regulating the adhesion and migration of both immune and tumor cells. Research has demonstrated that CTSZ enhances the infiltration of T cells through blood vessels, coordinating the tumor microenvironment in certain cancers .

CTSZ functions extend beyond basic cellular processes, as it participates in several immunological pathways and regulates cell-surface interactions critical for immune function. The protein interacts with cell surface integrins that mediate immune cell activity, including lymphocyte function-associated antigen-1 (LFA-1) and macrophage-1 (Mac-1) antigen, which regulates Mycobacterium tuberculosis (Mtb) phagocytosis and phagocyte migration .

How is CTSZ expression distributed across human tissues and cell types?

CTSZ demonstrates notable cell-type specificity in its expression pattern. Single-cell transcriptome data has revealed macrophage-specific expression of CTSZ in clear cell renal cell carcinoma (ccRCC) . Double immunofluorescence studies have confirmed that CTSZ is co-expressed with CD68 (a macrophage marker) but not with CD8 (a T cell marker) .

In tuberculosis research, CTSZ protein has been identified within CD68+ macrophages in human pulmonary granulomas, positioning it at the host-pathogen interface . Interestingly, in human-derived monocytes, CTSZ is highly expressed at baseline but becomes downregulated following Mtb infection . This dynamic expression pattern suggests context-dependent regulation that may influence disease outcomes.

What methods are recommended for detecting and quantifying CTSZ in experimental settings?

Multiple complementary approaches can be employed to detect and quantify CTSZ expression:

Transcript-level detection:

• RNA-Seq and HTSeq-FPKM/TPM format data analysis through databases like GEPIA and TCGA

• Single-cell transcriptome analysis for cell-type specific expression patterns

• RT-PCR with specific primers for detecting CTSZ variants

Protein-level detection:

• Immunohistochemistry (IHC) for tissue sections

• Double-labeling immunofluorescence (IF) using antibodies such as CTSZ (1:100, Santa Cruz, sc-376976) with cell-type markers like CD68 (1:1,000, Abcam, ab213363)

• Confocal microscopy visualization

Genetic analysis:

• Genotyping for CTSZ variants using PCR-based methods

• For wildtype CTSZ detection: forward primer 5'-TTG CTG TTG GCG AGT GCG-3' and reverse primer 5'-CTT GTC ACC AGA TTC CAG C-3'

These methodologies provide complementary information about CTSZ at multiple levels of biological organization, enabling comprehensive characterization in different research contexts.

How does CTSZ contribute to cancer progression and the tumor microenvironment?

CTSZ has emerged as a significant factor in multiple cancer types, including breast, colorectal, gastric, prostate, and hepatocellular carcinoma . In clear cell renal cell carcinoma (ccRCC), CTSZ is upregulated in tumor tissues compared to adjacent normal tissues at the RNA level, though this expression primarily occurs in tumor-infiltrating immune cells rather than in cancer cells themselves .

High CTSZ expression correlates with enrichment of several critical cancer pathways:

Clinically, CTSZ levels are associated with patient prognosis in ccRCC (hazard ratio=1.5, P=0.007), with patients having higher CTSZ expression showing worse outcomes with anti-PD-1 monotherapy (hazard ratio=1.51, P=0.039) . Some studies have proposed CTSZ as a putative oncogene, with increased expression associated with poor prognosis across multiple cancer types .

What is the relationship between CTSZ genetic variation and tuberculosis outcomes?

CTSZ has been identified as a conserved susceptibility factor in tuberculosis (TB) through both mouse models and human studies. In the Collaborative Cross mouse panel, which models human phenotypic and genotypic variation, cathepsin Z (Ctsz) emerged as a lead candidate underlying TB susceptibility .

In human studies from Uganda (n=328 across two independent cohorts), several CTSZ variants were significantly associated with TB disease severity after Bonferroni adjustment:

For rs113592645, the minor T allele was associated with both decreased TB disease severity and increased CTSZ expression following Mtb infection (p=0.0395) . This effect was specific to Mtb infection and not observed under mock infection conditions.

Mouse studies have demonstrated that Ctsz ablation leads to increased bacterial burden, CXCL1 overproduction, and decreased survival . These findings establish a CTSZ-CXCL1 axis that mediates TB disease severity in both humans and genetically diverse mice.

How does CTSZ expression influence immunotherapy response?

Research in clear cell renal cell carcinoma (ccRCC) has revealed important relationships between CTSZ expression and immunotherapy outcomes. High expression of CTSZ has been identified as a potential biomarker for predicting response to immunotherapy in ccRCC patients .

Specifically, patients with higher CTSZ expression demonstrated worse prognosis when treated with anti-PD-1 monotherapy (hazard ratio=1.51, P=0.039) . This correlation positions CTSZ as a potential treatment response biomarker that could guide clinical decision-making for patients receiving immune checkpoint inhibitor therapy.

The mechanistic basis for this relationship likely involves CTSZ's association with multiple immune checkpoint molecules, including CTLA4, LAG3, HAVCR2, PDCD1LG2, PDCD1, TIGIT, and SIGLEC15 . This immunoregulatory profile suggests that CTSZ may influence the tumor immune microenvironment in ways that affect response to immunotherapeutic interventions.

What is the CTSZ-CXCL1 axis and how does it influence inflammatory conditions?

The CTSZ-CXCL1 axis represents a newly identified immunoregulatory mechanism with particular relevance to tuberculosis and potentially other inflammatory conditions. Mouse studies have demonstrated that Ctsz ablation leads to consistent elevation of CXCL1 levels in the lungs both before and throughout Mtb infection .

At the cellular level, Ctsz−/− bone marrow-derived macrophages (BMDMs) produce significantly more CXCL1 in response to both pathogenic and non-pathogenic mycobacterial infection compared to wildtype cells . This finding suggests that CTSZ normally functions to regulate or suppress CXCL1 production in macrophages.

CXCL1 is a cytokine associated with severe TB disease in both mice and humans, primarily known for its role as a neutrophil chemoattractant . The overproduction of CXCL1 in the absence of CTSZ may contribute to excessive inflammation, neutrophil recruitment, and subsequent tissue damage during infection.

While the precise molecular mechanisms connecting CTSZ activity to CXCL1 regulation require further investigation, this axis represents an important target for understanding and potentially modulating inflammatory responses in infectious and other diseases.

How is CTSZ regulated at the epigenetic and transcriptional levels?

CTSZ expression appears to be regulated through multiple mechanisms, with epigenetic regulation playing a particularly important role:

DNA methylation:
Hypomethylation modification of specific CpG sites (cg02744249, cg02744249, and cg22145559) has been negatively correlated with CTSZ expression, suggesting an epigenetic regulatory mechanism . This relationship indicates that demethylation of these sites may enhance CTSZ transcription.

Genetic variant effects:
In a Ugandan cohort study, certain SNPs in the CTSZ gene were associated with divergent CTSZ transcription following Mtb infection. Most notably, the rs113592645 minor T allele was associated with increased CTSZ expression specifically in the context of Mtb infection (p=0.0395) .

Cell-type specific regulation:
CTSZ shows highly cell-type specific expression patterns, particularly in macrophages, suggesting lineage-specific transcriptional regulation. In human-derived monocytes, CTSZ is highly expressed at baseline but becomes downregulated following Mtb infection , indicating context-dependent transcriptional control.

These multi-level regulatory mechanisms likely contribute to the complex expression patterns of CTSZ across different tissues, cell types, and disease states.

What models are available for studying CTSZ function in human disease?

Researchers investigating CTSZ can employ several complementary model systems:

Genetically diverse mouse models:

• The Collaborative Cross (CC) mouse panel provides a powerful system for modeling human phenotypic and genotypic variation in CTSZ

• CC strains with the Tip5 S locus produce lower CTSZ protein levels and show higher bacterial burden following Mtb infection

Knockout models:

• Ctsz−/− mice allow investigation of complete CTSZ loss-of-function

• Targeted knockout in specific cell populations can dissect tissue-specific roles

Human genetic variation studies:

• Genome-wide association studies (GWAS) can identify CTSZ variants associated with disease outcomes

• The Bandim TBscore can quantify disease severity in relation to CTSZ variants

• Linkage disequilibrium analysis using tools like PLINK helps identify functional CTSZ haplotypes

Cell culture systems:

• Bone marrow-derived macrophages (BMDMs) from wildtype or Ctsz−/− mice

• Patient-derived monocytes with different CTSZ genotypes

• Cell lines with CTSZ knockdown or overexpression

Tissue-specific approaches:

• Human granuloma analysis for TB research

• Tumor tissue microarrays for cancer studies

These diverse models enable investigation of CTSZ function across different biological scales and disease contexts.

What techniques are recommended for analyzing CTSZ in human granulomas?

Studying CTSZ in human tuberculosis granulomas requires specialized approaches:

Tissue acquisition:

• Lung biopsies from patients with culture-confirmed TB, following ethical guidelines and obtaining appropriate consent

• Careful preservation of tissue architecture for subsequent analysis

Immunostaining approaches:

• Double-labeling immunofluorescence using CTSZ antibodies (1:100, Santa Cruz, sc-376976) alongside macrophage markers like CD68 (1:1,000, Abcam, ab213363)

• Confocal microscopy visualization to precisely localize CTSZ within specific granuloma cell populations

• Serial section analysis to understand spatial distribution throughout the granuloma structure

Advanced methods:

• Single-cell RNA sequencing of dissociated granuloma tissue

• Spatial transcriptomics to preserve positional information

• Multiplexed immunofluorescence for simultaneous detection of multiple markers

These approaches have revealed that CTSZ is produced within CD68+ macrophages in human TB granulomas, positioning it at the host-pathogen interface . This localization suggests that CTSZ may play important roles in granuloma function and Mtb containment.

How can single-cell transcriptomics advance our understanding of CTSZ function?

Single-cell transcriptomics offers powerful insights into CTSZ biology across different cell types and disease states:

Cell type-specific expression mapping:

• Single-cell RNA sequencing has revealed macrophage-specific expression of CTSZ in ccRCC and other contexts

• This cell type specificity explains why CTSZ is upregulated in some tumor tissues but not in tumor cells themselves

Co-expression network analysis:

• Identification of genes whose expression patterns correlate with CTSZ in specific cell populations

• Inference of functional relationships and signaling pathways through computational approaches

Heterogeneity characterization:

• Even within macrophage populations, identification of subpopulations with different CTSZ expression levels

• Correlation of this heterogeneity with disease outcomes and treatment responses

Integration with genetic data:

• Analysis of how CTSZ genetic variants affect expression in specific cell types

• Mechanistic insights into disease-associated polymorphisms

Analytical approaches:

• Software packages like "DESeq2", "ggplot2", "clusterProfiler", and "org.Hs.e.g.db" for analyzing CTSZ-correlated genes

• Gene Ontology and KEGG pathway analyses to identify enriched biological processes

By leveraging these approaches, researchers can gain unprecedented resolution in understanding CTSZ function within the complex cellular ecosystems of diseases like cancer and tuberculosis.

What are the conflicting findings regarding CTSZ's role in disease prognosis?

The literature presents some apparently contradictory findings regarding CTSZ's impact on disease outcomes:

In cancer research:

• Increased CTSZ expression is associated with poor patient prognoses in several cancer types, with some studies proposing CTSZ as a putative oncogene

• In ccRCC, high CTSZ expression correlates with worse prognosis (hazard ratio=1.5, P=0.007)

• Patients with higher CTSZ expression show worse outcomes with anti-PD-1 monotherapy (hazard ratio=1.51, P=0.039)

In tuberculosis research:

• The rs113592645 minor T allele is associated with decreased TB disease severity

• This same allele correlates with increased CTSZ expression following Mtb infection

• Conversely, mouse studies demonstrate that complete Ctsz ablation leads to increased bacterial burden and decreased survival

These findings suggest a complex, context-dependent relationship between CTSZ expression and disease outcomes that may depend on:

• Disease type (cancer vs. infectious disease)

• Cell-specific expression patterns

• Interaction with other pathways (e.g., CXCL1 axis in TB, immune checkpoint pathways in cancer)

• Degree of expression change (partial vs. complete loss)

The data indicate that balanced CTSZ levels may be required for optimal outcomes, with both excessive and insufficient expression potentially contributing to disease progression in different contexts.

What knowledge gaps remain in understanding CTSZ's physiological functions?

Despite significant progress in CTSZ research, several important knowledge gaps remain:

Molecular mechanism of CTSZ-CXCL1 regulation:
While the relationship between CTSZ and CXCL1 has been established in TB models, the precise molecular mechanism connecting these factors requires further investigation .

Cell type-specific functions:
Although CTSZ shows macrophage-specific expression in certain contexts, its functions in other cell types and tissues remain incompletely characterized.

Interaction with immune checkpoints:
The consistent correlation between CTSZ and immune checkpoint molecules suggests functional relationships, but the mechanistic basis needs clarification .

Therapeutic potential:
Despite associations with disease outcomes, the potential for targeting CTSZ therapeutically has not been thoroughly explored.

Regulation of CTSZ itself:
While some epigenetic mechanisms have been identified, the comprehensive regulatory landscape controlling CTSZ expression across different contexts remains to be elucidated.

Addressing these knowledge gaps will require integrated approaches combining genetic, molecular, cellular, and clinical investigations to fully understand CTSZ's multifaceted roles in human biology and disease.