The research report should be a detailed narrative explaining the function, biological processes, and localization of the gene product. Citations should be given for all claims.
You should prioritize authoritative reviews and primary scientific literature when conducting research. You can supplement
this with annotations you find in gene/protein databases, but these can be outdated or inaccurate.
We are specifically interested in the primary function of the gene - for enzymes, what reaction is catalyzed, and what is the substrate specificity? For transporters, what is the substrate? For structural proteins or adapters, what is the broader structural role? For signaling molecules, what is the role in the pathway.
We are interested in where in or outside the cell the gene product carries out its function.
We are also interested in the signaling or biochemical pathways in which the gene functions. We are less interested in broad pleiotropic effects, except where these elucidate the precise role.
Include evidence where possible. We are interested in both experimental evidence as well as inference from structure, evolution, or bioinformatic analysis. Precise studies should be prioritized over high-throughput, where available.
The requested protein is correctly identified. In Schizosaccharomyces pombe, cis4 corresponds to ORF SPAC17D4.03c and encodes a 732-amino-acid cation diffusion facilitator (CDF), related to fungal Msc2 and metazoan ZnT5/6/7 proteins. The identification was established by complementation, sequencing, and linkage analysis of 30 tetrads—not merely by name similarity. This matches the supplied UniProt accession Q9HGQ3, organism, and Msc2-like/Cation_efflux domain annotations. No literature concerning a different same-named gene was used. (fang2008cationdiffusionfacilitator pages 1-2, fang2008cationdiffusionfacilitator pages 2-4)
The best-supported functional annotation is:
Cis4 is the membrane component of a Cis4–Zrg17 heteromeric zinc-transport complex in the cis-Golgi. It removes Zn²⁺ from the cytosol—particularly during zinc limitation—and supplies it to the Golgi/early secretory-pathway lumen, thereby supporting zinc-dependent secretory functions and Golgi membrane trafficking.
Zn²⁺ specificity is strongly supported physiologically and by cellular zinc measurements. However, transport has not been reconstituted with purified Cis4–Zrg17, so its affinity, coupling ion, stoichiometry, exact selectivity spectrum, and catalytic transport cycle remain unknown. (fang2008cationdiffusionfacilitator pages 4-5, choi2018zinctransportersbelonging pages 13-16, choi2018zinctransportersbelonging pages 3-6)
| Annotation | Conclusion | Evidence type | Confidence / limitations |
|---|---|---|---|
| Identity | cis4 is the Schizosaccharomyces pombe gene SPAC17D4.03c, encoding a 732-aa protein; linkage analysis of 30 tetrads associated this ORF with the cis4 locus. This matches the supplied UniProt accession Q9HGQ3 and organism context. (fang2008cationdiffusionfacilitator pages 1-2, fang2008cationdiffusionfacilitator pages 2-4) | Direct genetic mapping, complementation, and sequencing | High. The experimental gene designation and ORF match the requested target; the UniProt accession itself was supplied by the user rather than independently established in these studies. |
| Family and domains | Cis4 is a cation diffusion facilitator (CDF) protein homologous to budding-yeast Msc2 and mammalian ZnT5/6/7. The specific Cation_efflux, Cation_efflux_TM, Cation_efflux_TMD_sf, Msc2-like, and PF01545 assignments are supplied UniProt/InterPro/Pfam annotations, not experimentally mapped Cis4 domains. (fang2008cationdiffusionfacilitator pages 2-4, fang2008cationdiffusionfacilitator pages 7-8) | Sequence homology and database annotation | High for CDF/Msc2-like family membership; moderate for exact domain boundaries and topology because no Cis4 structure or domain-mapping experiment was identified. |
| Substrate and direction | The best-supported substrate is Zn²⁺. The Cis4–Zrg17 complex removes zinc from the cytosol and is modeled to import it into the cis-Golgi/secretory-pathway lumen. Zinc-specific phenotypic rescue and cytosolic FRET measurements support this assignment, but transport across purified membranes has not been directly measured. (fang2008cationdiffusionfacilitator pages 4-5, choi2018zinctransportersbelonging pages 13-16, choi2018zinctransportersbelonging pages 3-6) | Metal-specific rescue, genetically encoded cytosolic-zinc sensors, genetics, and localization-based inference | High for zinc as the physiological substrate; moderate-to-high for cytosol-to-Golgi direction. Direct kinetics, coupling ion, stoichiometry, affinity, and broader metal selectivity remain unknown. |
| Localization | Functional Cis4-RFP occurs in cytoplasmic puncta that colocalize with the cis-Golgi marker Rer1-GFP, but not the trans-Golgi marker Krp1-GFP. (fang2008cationdiffusionfacilitator pages 4-5) | Direct fluorescence colocalization using a functional tagged protein | High for cis-Golgi localization under the tested conditions. Membrane orientation and possible minor localization elsewhere were not resolved. |
| Functional partner | Cis4 functions with the CDF protein Zrg17 as a heteromeric complex. GST pulldown detected GFP-Cis4 with GST-Zrg17, and the two mutants showed similar, nonadditive, zinc-suppressible phenotypes; coexpression of budding-yeast MSC2 and ZRG17 rescued cis4 defects. (fang2008cationdiffusionfacilitator pages 1-2, fang2008cationdiffusionfacilitator pages 4-5) | Direct biochemical interaction, genetic epistasis, and heterologous complementation | High. Complex stoichiometry, assembly mechanism, and the respective transport roles of each subunit have not been established. |
| Physiological condition | Cis4–Zrg17 is most important for exporting zinc from the cytosol during zinc limitation, whereas ER-localized Zhf1 dominates when labile zinc is abundant. cis4Δ and zrg17Δ increase cytosolic zinc-sensor saturation during deficiency; their transcripts are not detectably regulated by zinc. Estimated protein abundance was approximately 1,300–5,500 Cis4, 3,000–6,000 Zrg17, and 12,000–14,000 Zhf1 molecules per cell. (choi2018zinctransportersbelonging pages 13-16, choi2018zinctransportersbelonging pages 16-18) | FRET zinc sensing, mutant analysis, transcript assays, and published proteomic estimates | Moderate-to-high. Condition-specific transport activity is supported, but its biochemical basis may involve affinity, abundance, localization, or post-translational control and remains unresolved. |
| Trafficking phenotypes | cis4 mutants have weakened cell-wall integrity and defective secretion. Acid-phosphatase secretion and total accumulation fell to approximately 25% and 55% of wild type, respectively. cis4 deletion also produced synthetic growth defects with ypt3-i5, ryh1-i6, gdi1-i11, and apm1-1 trafficking mutants; zinc rescued selected defects. (fang2008cationdiffusionfacilitator pages 1-2, fang2008cationdiffusionfacilitator pages 4-5, fang2008cationdiffusionfacilitator pages 7-8) | Quantitative secretion assays, cell-wall assays, genetic interactions, and zinc rescue | High that Cis4-dependent zinc homeostasis affects Golgi trafficking; moderate for the precise mechanism because the relevant zinc-requiring Golgi clients are unidentified. |
| GPI-anchored-protein link | Multicopy ecm33⁺, aah3⁺, and gaz2⁺, encoding GPI-anchored proteins, suppressed the MgCl₂-sensitive cis4-1 phenotype; cis4 mutants were also sensitive to the GPI-synthesis inhibitor BE49385A. However, GFP-Ecm33 localization was normal in cis4Δ cells under standard conditions, arguing against a simple primary block in GPI-anchor synthesis or ER export. (jaiseng2012studiesonthe pages 7-8, jaiseng2012studiesonthe pages 1-2) | Multicopy suppression, inhibitor sensitivity, genetic interaction, and fluorescence localization | Moderate. Cis4 is linked indirectly to GPI-protein trafficking or cell-surface function, but it is not established as a dedicated GPI-trafficking factor. |
| Principal uncertainties | No Cis4 atomic structure, purified-protein transport assay, transport kinetics, coupling mechanism, membrane orientation, exact selectivity series, or identified zinc-dependent Golgi client was found. The designation “probable zinc transporter” is therefore strongest at the physiological level rather than as a fully reconstituted biochemical mechanism. (fang2008cationdiffusionfacilitator pages 4-5, choi2018zinctransportersbelonging pages 13-16, fang2008cationdiffusionfacilitator pages 7-8) | Evidence-gap assessment | Important limitation. Current annotation combines strong genetic, localization, interaction, and cellular-zinc evidence with family-based mechanistic inference. |
Table: This table summarizes the identity, molecular function, localization, physiological role, and evidence gaps for S. pombe cis4/Q9HGQ3. It distinguishes direct experimental findings from database annotation and family-based inference.
The foundational study cloned SPAC17D4.03c by complementation of the MgCl₂- and FK506-sensitive cis4-1 mutant. Linkage analysis of 30 tetrads yielded only parental ditypes, identifying SPAC17D4.03c as cis4. Sequencing predicted a 732-aa CDF-family protein; cis4-1 and cis4-2 carried Gly→Glu at residue 375 and Gly→Asp at residue 393, respectively. These data decisively connect the historical genetic locus, ORF, and protein discussed in the literature. (fang2008cationdiffusionfacilitator pages 1-2, fang2008cationdiffusionfacilitator pages 2-4)
The supplied database assignments—Cation_efflux/IPR002524, Cation_efflux_TM/IPR058533, Cation_efflux_TMD superfamily/IPR027469, Msc2-like/IPR045316, and PF01545—are consistent with this experimentally established CDF/Msc2-like identity. Exact domain boundaries and membrane topology should nevertheless be treated as computational annotations because no Cis4-specific structure or experimental topology map was found. (fang2008cationdiffusionfacilitator pages 2-4, fang2008cationdiffusionfacilitator pages 7-8)
Several independent observations identify zinc as the physiological substrate:
Collectively, these results provide high confidence that Cis4 transports Zn²⁺ physiologically. They do not exclude low-level transport of another metal, but no direct evidence establishes an alternative substrate.
The current model is cytosol → cis-Golgi/secretory-pathway lumen. This direction follows from elevated cytosolic zinc after cis4 deletion, the complex’s cis-Golgi localization, and the general outward-from-cytosol activity of the relevant CDF group. The direction is strongly supported at the cellular level but was not measured directly across purified Golgi membranes. (fang2008cationdiffusionfacilitator pages 4-5, choi2018zinctransportersbelonging pages 13-16, choi2018zinctransportersbelonging pages 3-6)
Cis4 is therefore not primarily a plasma-membrane zinc importer. It is an intracellular zinc-distribution transporter, converting cytosolic zinc availability into luminal zinc availability for the early secretory pathway.
A functional Cis4-RFP fusion appeared in punctate cytoplasmic structures and colocalized with Rer1-GFP, a cis-Golgi marker, but not with the trans-Golgi marker Krp1-GFP. The relevant site of action is consequently the cis-Golgi membrane. Membrane orientation and whether a minor pool occurs in other secretory compartments remain unresolved. (fang2008cationdiffusionfacilitator pages 4-5)
Cis4 functions with another CDF protein, Zrg17. GST pulldown recovered GFP-Cis4 with GST-Zrg17 but not GST alone, directly demonstrating association. Similar, nonadditive, zinc-rescuable mutant phenotypes additionally place both proteins in the same functional unit. The literature consequently describes the transporter as a Cis4/Zrg17 heteromer, although its exact subunit stoichiometry and each subunit’s mechanistic contribution are unknown. (fang2008cationdiffusionfacilitator pages 1-2, fang2008cationdiffusionfacilitator pages 4-5)
Cis4–Zrg17 and the ER-localized CDF transporter Zhf1 partition zinc out of the cytosol in a condition-dependent manner. During zinc deficiency, deleting cis4 or zrg17 increases cytosolic ZapCY1 saturation, indicating that Cis4–Zrg17 remains important for secretory-pathway zinc allocation even when zinc is scarce. Zhf1 instead dominates cytosolic zinc removal following zinc exposure or under zinc-replete conditions. (choi2018zinctransportersbelonging pages 12-13, choi2018zinctransportersbelonging pages 2-3, choi2018zinctransportersbelonging pages 13-16)
This specialization is not explained by zinc-responsive transcription: cis4, zrg17, and zhf1 transcript abundance was reported to be independent of zinc status and the Loz1 zinc-responsive regulator. Proposed explanations include different zinc affinities, abundance, localization, or post-translational control, but these mechanisms have not been resolved. Published abundance estimates are approximately 1,300–5,500 Cis4, 3,000–6,000 Zrg17, and 12,000–14,000 Zhf1 molecules per cell, potentially contributing to Zhf1’s dominance when zinc is plentiful. (choi2018zinctransportersbelonging pages 13-16, choi2018zinctransportersbelonging pages 16-18)
Loss of Cis4 causes cell-wall and secretory defects. In cis4-deficient cells, secreted acid phosphatase fell to approximately 25% of wild type, while total cellular accumulation was about 55% of wild type. The disproportionate secretion defect implicates membrane trafficking rather than transcription alone. cis4 deletion also caused synthetic growth defects with the trafficking alleles ypt3-i5, ryh1-i6, gdi1-i11, and apm1-1. Zinc supplementation rescued selected trafficking phenotypes, notably those of ypt3-i5 and apm1-1, reinforcing a mechanistic connection between luminal zinc homeostasis and Golgi function. (fang2008cationdiffusionfacilitator pages 1-2, fang2008cationdiffusionfacilitator pages 4-5, fang2008cationdiffusionfacilitator pages 7-8)
The most defensible pathway interpretation is that Cis4 does not act as a coat, tether, or trafficking adaptor. Rather, it supplies Golgi zinc required by one or more unidentified luminal zinc-dependent clients whose maturation or activity supports vesicle trafficking, glycoprotein processing, secretion, or cell-wall construction. The specific client proteins remain unknown. (fang2008cationdiffusionfacilitator pages 7-8)
Multicopy expression of three GPI-anchored proteins—Ecm33, Aah3, and Gaz2—suppressed the cis4-1 MgCl₂-sensitive phenotype; Aah3 and Gaz2 gave stronger suppression than Ecm33. cis4 mutants were also sensitive to BE49385A, an inhibitor of the GPI-biosynthetic protein Its8. These genetic observations connect Cis4-dependent Golgi function with GPI-anchored proteins and cell-surface integrity. (jaiseng2012studiesonthe pages 7-8, jaiseng2012studiesonthe pages 1-2)
This connection is probably indirect. GFP-Ecm33 localization was normal in cis4Δ cells under standard conditions, with no abnormal ER accumulation. Thus, present evidence does not support annotating Cis4 as a dedicated GPI-anchor biosynthesis or GPI-cargo transporter. (jaiseng2012studiesonthe pages 7-8)
cis4 deletion is viable, indicating that Cis4 is not essential under routine laboratory growth conditions. Mutants are sensitive to MgCl₂ and FK506 and exhibit weakened cell-wall integrity, reduced acid-phosphatase secretion, and genetic interactions with membrane-trafficking mutants. The original cis4-1 allele was synthetically lethal with calcineurin deletion, although allele-specific behavior complicates interpretation: cis4Δ was less FK506-sensitive than cis4-1. These broad stress phenotypes are best regarded as downstream consequences of disturbed Golgi zinc homeostasis rather than evidence that Cis4 directly transports magnesium or participates in calcineurin signaling. (fang2008cationdiffusionfacilitator pages 7-8, fang2008cationdiffusionfacilitator pages 2-4)
Under zinc-limited conditions containing 100 μM EDTA, cis4Δ shows a slight growth defect, while zrg17Δ is somewhat more sensitive. Growth was assessed across 0–200 μM added zinc, with plates incubated for 3–5 days at 31°C. More importantly, FRET sensors directly detected abnormal cytosolic zinc partitioning in these mutants, moving the annotation beyond inference from growth alone. (choi2018zinctransportersbelonging pages 13-16, choi2018zinctransportersbelonging pages 3-6)
The Cis4-specific evidence base is sparse. Searches identified a 2023 review of zinc homeostasis in S. pombe and a 2024 dissertation concerning zinc-responsive proteins, but no accessible 2023–2024 primary study that directly revised Cis4’s substrate, localization, or mechanism. Consequently, the newest decisive Cis4-specific primary evidence remains the 2018 PLOS Genetics FRET study. It refined the older model by demonstrating that Cis4–Zrg17 and Zhf1 have complementary, zinc-condition-dependent roles rather than simply redundant functions. (choi2018zinctransportersbelonging pages 13-16, choi2018zinctransportersbelonging pages 12-13)
A 2023 review on secretory-pathway Zn²⁺ enzyme metalation places transporters of this class in a broader current framework: luminal zinc delivery is required for maturation of zinc-dependent secretory proteins. For Cis4, however, a particular Golgi zinc enzyme or client has not yet been identified; application of the general model to Cis4 remains mechanistic inference rather than direct evidence.
Relevant recent and authoritative URLs include:
Cis4 currently has no direct clinical or industrial implementation. Its practical value is as a genetically tractable model for:
Such cross-species extrapolation should remain cautious: homology to mammalian ZnT proteins does not establish identical substrate kinetics, topology, regulation, or client proteins in Cis4.
No study retrieved here provides a Cis4 atomic structure, purified-protein transport assay, Zn²⁺ affinity, turnover rate, coupling-ion identity, transport stoichiometry, experimentally resolved topology, or comprehensive metal-selectivity series. No specific zinc-dependent Golgi enzyme has been shown to mediate the trafficking phenotype.
Accordingly, a precise annotation would be:
“Cis-Golgi-localized component of the heteromeric Cis4–Zrg17 CDF zinc transporter; mediates transfer of cytosolic Zn²⁺ into the Golgi/early secretory pathway, particularly under zinc limitation, and thereby supports Golgi membrane trafficking, secretion, and cell-wall homeostasis.”
Confidence is high for identity, CDF-family membership, Zrg17 interaction, cis-Golgi localization, and physiological Zn²⁺ transport; moderate-to-high for cytosol-to-Golgi direction; and low/unknown for biochemical kinetics, coupling mechanism, exact topology, and direct downstream zinc clients. The foundational primary reports were published in April 2008 (https://doi.org/10.1091/mbc.e07-08-0805), July 2012 (https://doi.org/10.1371/journal.pone.0041946), and March 2018 (https://doi.org/10.1371/journal.pgen.1007262). (fang2008cationdiffusionfacilitator pages 1-2, fang2008cationdiffusionfacilitator pages 4-5, choi2018zinctransportersbelonging pages 13-16, jaiseng2012studiesonthe pages 7-8)
References
(fang2008cationdiffusionfacilitator pages 1-2): Yue Fang, R. Sugiura, Yan Ma, Tomoko Yada-Matsushima, H. Umeno, and T. Kuno. Cation diffusion facilitator cis4 is implicated in golgi membrane trafficking via regulating zinc homeostasis in fission yeast. Molecular biology of the cell, 19 4:1295-303, Apr 2008. URL: https://doi.org/10.1091/mbc.e07-08-0805, doi:10.1091/mbc.e07-08-0805. This article has 41 citations and is from a domain leading peer-reviewed journal.
(fang2008cationdiffusionfacilitator pages 2-4): Yue Fang, R. Sugiura, Yan Ma, Tomoko Yada-Matsushima, H. Umeno, and T. Kuno. Cation diffusion facilitator cis4 is implicated in golgi membrane trafficking via regulating zinc homeostasis in fission yeast. Molecular biology of the cell, 19 4:1295-303, Apr 2008. URL: https://doi.org/10.1091/mbc.e07-08-0805, doi:10.1091/mbc.e07-08-0805. This article has 41 citations and is from a domain leading peer-reviewed journal.
(fang2008cationdiffusionfacilitator pages 4-5): Yue Fang, R. Sugiura, Yan Ma, Tomoko Yada-Matsushima, H. Umeno, and T. Kuno. Cation diffusion facilitator cis4 is implicated in golgi membrane trafficking via regulating zinc homeostasis in fission yeast. Molecular biology of the cell, 19 4:1295-303, Apr 2008. URL: https://doi.org/10.1091/mbc.e07-08-0805, doi:10.1091/mbc.e07-08-0805. This article has 41 citations and is from a domain leading peer-reviewed journal.
(choi2018zinctransportersbelonging pages 13-16): Sangyong Choi, Ya-Mei Hu, Mark E. Corkins, Amy E. Palmer, and Amanda J. Bird. Zinc transporters belonging to the cation diffusion facilitator (cdf) family have complementary roles in transporting zinc out of the cytosol. Mar 2018. URL: https://doi.org/10.1371/journal.pgen.1007262, doi:10.1371/journal.pgen.1007262. This article has 45 citations and is from a domain leading peer-reviewed journal.
(choi2018zinctransportersbelonging pages 3-6): Sangyong Choi, Ya-Mei Hu, Mark E. Corkins, Amy E. Palmer, and Amanda J. Bird. Zinc transporters belonging to the cation diffusion facilitator (cdf) family have complementary roles in transporting zinc out of the cytosol. Mar 2018. URL: https://doi.org/10.1371/journal.pgen.1007262, doi:10.1371/journal.pgen.1007262. This article has 45 citations and is from a domain leading peer-reviewed journal.
(fang2008cationdiffusionfacilitator pages 7-8): Yue Fang, R. Sugiura, Yan Ma, Tomoko Yada-Matsushima, H. Umeno, and T. Kuno. Cation diffusion facilitator cis4 is implicated in golgi membrane trafficking via regulating zinc homeostasis in fission yeast. Molecular biology of the cell, 19 4:1295-303, Apr 2008. URL: https://doi.org/10.1091/mbc.e07-08-0805, doi:10.1091/mbc.e07-08-0805. This article has 41 citations and is from a domain leading peer-reviewed journal.
(choi2018zinctransportersbelonging pages 16-18): Sangyong Choi, Ya-Mei Hu, Mark E. Corkins, Amy E. Palmer, and Amanda J. Bird. Zinc transporters belonging to the cation diffusion facilitator (cdf) family have complementary roles in transporting zinc out of the cytosol. Mar 2018. URL: https://doi.org/10.1371/journal.pgen.1007262, doi:10.1371/journal.pgen.1007262. This article has 45 citations and is from a domain leading peer-reviewed journal.
(jaiseng2012studiesonthe pages 7-8): Wurentuya Jaiseng, Yue Fang, Yan Ma, Reiko Sugiura, and Takayoshi Kuno. Studies on the roles of clathrin-mediated membrane trafficking and zinc transporter cis4 in the transport of gpi-anchored proteins in fission yeast. PLoS ONE, 7:e41946, Jul 2012. URL: https://doi.org/10.1371/journal.pone.0041946, doi:10.1371/journal.pone.0041946. This article has 12 citations and is from a peer-reviewed journal.
(jaiseng2012studiesonthe pages 1-2): Wurentuya Jaiseng, Yue Fang, Yan Ma, Reiko Sugiura, and Takayoshi Kuno. Studies on the roles of clathrin-mediated membrane trafficking and zinc transporter cis4 in the transport of gpi-anchored proteins in fission yeast. PLoS ONE, 7:e41946, Jul 2012. URL: https://doi.org/10.1371/journal.pone.0041946, doi:10.1371/journal.pone.0041946. This article has 12 citations and is from a peer-reviewed journal.
(choi2018zinctransportersbelonging pages 12-13): Sangyong Choi, Ya-Mei Hu, Mark E. Corkins, Amy E. Palmer, and Amanda J. Bird. Zinc transporters belonging to the cation diffusion facilitator (cdf) family have complementary roles in transporting zinc out of the cytosol. Mar 2018. URL: https://doi.org/10.1371/journal.pgen.1007262, doi:10.1371/journal.pgen.1007262. This article has 45 citations and is from a domain leading peer-reviewed journal.
(choi2018zinctransportersbelonging pages 2-3): Sangyong Choi, Ya-Mei Hu, Mark E. Corkins, Amy E. Palmer, and Amanda J. Bird. Zinc transporters belonging to the cation diffusion facilitator (cdf) family have complementary roles in transporting zinc out of the cytosol. Mar 2018. URL: https://doi.org/10.1371/journal.pgen.1007262, doi:10.1371/journal.pgen.1007262. This article has 45 citations and is from a domain leading peer-reviewed journal.