Die Isolierung und Aufreinigung der Metalloendopeptidase LysN und die Evaluierung der Spezifität der LysN E157D-Mutante und ihr Einsatz in der Proteomik und die Mechanismen der Bildung von “dendritic cell aggresome- like induced structures“ (DALIS)

Der proteolytische Verdau von Proteinen in Peptide ist ein wichtiger Schritt in der Tandem-Massenspektrometrie. Dabei werden Peptide fragmentiert und die sich ergebenden Fragmentionen geben Aufschluss über die Aminosäuresequenz des zu untersuchenden Proteins. Dabei sind für die Fragmentierung sowohl Länge und Sequenz, als auch der Ladungszustand des Peptids ungemein wichtig. Diese Parameter bedingen sich durch Endoproteasen, die für den proteolytischen Verdau eingesetzt werden. Eine Voraussetzung hierfür ist die Spezifität der Protease. Trypsin ist bei weitem die gebräuchlichste Protease zur massenspektrometrischen Probenvorbereitung. Allerdings bietet Trypsin keine Komplettlösung. Je nach Fragestellung und Applikation müssen weitere Proteasen eingesetzt werden, um eine komplette Sequenzabdeckung zu gewährleisten und möglichst alle posttranslationalen Modifikationen nachzuweisen, oder bestimmte Proteomklassen (z.B Phosphoproteom

Functional characterization of the placenta specific protein PLAC1 and its use for cancer immunotherapy

Summary Antibody-based cancer therapies have been successfully introduced into the clinic and have emerged as the most promising therapeutics in oncology. The limiting factor regarding the development of therapeutical antibody vaccines is the identification of tumor-associated antigens. PLAC1, the placenta-specific protein 1, was categorized for the first time by the group of Prof. Sahin as such a tumor-specific antigen. Within this work PLAC1 was characterized using a variety of biochemical methods. The protein expression profile, the cellular localization, the conformational state and especially the interacting partners of PLAC1 and its functionality in cancer were analyzed. Analysis of the protein expression profile of PLAC1 in normal human tissue confirms the published RT-PCR data. Except for placenta no PLAC1 expression was detectable in any other normal human tissue. Beyond, an increased PLAC1 expression was detected in several cancer cell lines derived of trophoblastic, breast and pancreatic lineage emphasizing its properties as tumor-specific antigen. rnThe cellular localization of PLAC1 revealed that PLAC1 contains a functional signal peptide which conducts the propeptide to the endoplasmic reticulum (ER) and results in the secretion of PLAC1 by the secretory pathway. Although PLAC1 did not exhibit a distinct transmembrane domain, no unbound protein was detectable in the cell culture supernatant of overexpressing cells. But by selective isolation of different cellular compartments PLAC1 was clearly enriched within the membrane fraction. Using size exclusion chromatography PLAC1 was characterized as a highly aggregating protein that forms a network of high molecular multimers, consisting of a mixture of non-covalent as well as covalent interactions. Those interactions were formed by PLAC1 with itself and probably other cellular components and proteins. Consequently, PLAC1 localize outside the cell, where it is associated to the membrane forming a stable extracellular coat-like structure.rnThe first mechanistic hint how PLAC1 promote cancer cell proliferation was achieved identifying the fibroblast growth factor FGF7 as a specific interacting partner of PLAC1. Moreover, it was clearly shown that PLAC1 as well as FGF7 bind to heparin, a glycosaminoglycan of the ECM that is also involved in FGF-signaling. The participation of PLAC1 within this pathway was approved after co-localizing PLAC1, FGF7 and the FGF7 specific receptor (FGFR2IIIb) and identifying the formation of a trimeric complex (PLAC1, FGF7 and the specific receptor FGFR2IIIb). Especially this trimeric complex revealed the role of PLAC1. Binding of PLAC1 together with FGF7 leads to the activation of the intracellular tyrosine kinase of the FGFR2IIIb-receptor and mediate the direct phosphorylation of the AKT-kinase. In the absence of PLAC1, no FGF7 mediated phosphorylation of AKT was observed. Consequently the function of PLAC1 was clarified: PLAC1 acts as a co-factor by stimulating proliferation by of the FGF7-FGFR2 signaling pathway.rnAll together, these novel biochemical findings underline that the placenta specific protein PLAC1 could be a new target for cancer immunotherapy, especially considering its potential applicability for antibody therapy in tumor patients.

Chaperone BiP: Master regulator of the unfolded protein response (UPR) and its role in regulation of angiogenesis

Verschiedene Krankheiten gehen mit einer fehlerhaften Vaskularisierung einher. Allerdings ist der Erfolg der derzeitig vorhandenen Therapieansätze, die sich z.B. auf VEGF fokussieren, beschränkt. Aus diesem Grund ist es wichtig, neue Strategien zur Regulation der Angiogenese zu entwickeln. Hierbei stehen neue Signaltransduktions-wege im Fokus, die sich als vielversprechend erweisen, um Angiogenese zu fördern oder zu inhibieren. Die Blutgefäßneubildung ist ein hochregulierter Prozess, der mit einer hohen Proteinsyntheserate verknüpft ist. Die Angiogenese wurde bereits mit dem ER-Stress Signaltransduktionsweg, der Unfolded Protein Response (UPR), in Verbindung gebracht (Zeng et al., 2013; Bouvier et al., 2012). Eine im Rahmen der vorliegenden Studie durchgeführte histologische Untersuchung konnte eine Fehlregulierung der Expression von UPR beteiligten Proteinen in vivo unter pathologischen Bedingungen gezeigt werden. Bemerkenswerter Weise war BiP, der Hauptsensor der UPR, in Endothelzellen von Angiosarkomen sehr stark exprimiert. In in vitro Experimenten wurde gezeigt, dass das Herunterregulieren von BiP mittels RNAi Einfluss auf die inflammatorische Antwort und die Bildung angiogener Strukturen in Endothelzellen nimmt. Das Herunterregulieren des Proteins BiP verstärkte die inflammatorische Antwort von HUVEC, was sich in einer gesteigerten Bildung von IL-8 und ICAM-1 äußerte und wurde auf die Aktivierung der UPR durch die verringerte Menge an BiP zurückgeführt. Der Phänotyp BiP-herunterregulierter Zellen entsprach dem untransfizierter Zellen, welcher durch das Cytoskelett und die Expression des endothelspezifischen Markers CD31 charakterisiert wurde. Im Gegensatz dazu änderte sich der Grad der Glykosylierung in transfizierten Zellen. Im Hinblick auf die Blutgefäßbildung, zeigten sich eine gehemmte Migration und eine inhibierte Bildung Gefäß-ähnlicher Strukturen in BiP-herunterregulierten Zellen. In diesen Zellen war die Expression von KDR auffallend stark inhibiert, wohingegen die Flt-1 Expression sich als gleichbleibend herausstellte, was ebenfalls auf die Aktivierung der UPR zurückgeführt werden konnte. Alternativ wäre der reduzierte Level des Proteins BiP im Hinblick auf die Funktion als Helferenzym in der Proteinfaltung eine mögliche Erklärung für die gehemmte Expression von KDR. Die Ergebnisse dieser Studie deuten darauf hin, dass stabile Spiegel von BiP die Regulierung der Angiogenese durch die Kontrolle der UPR in physiologischen Prozessen unterstützen könnte. Eine Fehlregulierung von BiP durch Unterdrückung der UPR, wie z.B. in malignen Tumoren, könnte Tumorzellen und beteiligten Endothelzellen einen Vorteil verschaffen und zu einer gestörten Vaskularisierung führen. Somit stellt das Stresssensorprotein BiP und die UPR einen potentiellen Angriffspunkt für die Regulation der Angiogenese dar.

Leishmania major promastigote entry of an autophagy-like compartment and amastigote escape from the parasitophorous vacuole

In this thesis, we investigated the interaction of the obligate intracellular parasite Leishmania (L.) major with two phenotypes of human monocyte derived macrophages (hMDMs). Thereby we focused on the development and maturation of the parasitophorous vacuole (PV) and could show that compartment development is dependent on the parasite stage.rnFocusing on the ultrastructure of PVs containing axenic amastigotes, we demonstrated that the parasites are partially located in damaged PVs or in the cytoplasm of the host. Moreover, we visualized multiple amastigotes in a common PV 144 h p.i. in pro-inflammatory hMDM I but not in anti-inflammatory hMDM II indicating different PV development. rnRegarding the promastigote form, we demonstrated a different uptake of viable and apoptotic L. major promastigotes by hMDMs. Viable promastigotes are predominantly taken up via the flagellum tip whereas apoptotic promastigotes enter the cells via the parasite body. Analyzing compartment maturation, we found that 20-30% of the PVs get positive for the early maturation markers PI3P and EEA1 independent of the viability of the parasites and unaffected by the human macrophage type. Subsequently, 25-40% of the parasites acquire the autophagy marker LC3 on their PV, what is independent of the viability of the parasites as well. We quantified this and in hMDM II less LC3-positive compartments formed compared to hMDM I. Analyzing the ultrastructure, we investigated that the compartments consist of a single-membrane PV characteristic for LC3-associated phagocytosis (LAP). Involvement of LAP was confirmed by demonstrating that the protein kinase ULK1 is dispensable for LC3-compartment formation around Leishmania PVs. Visualizing compartment dynamics in real time showed that apoptotic promastigotes are degraded in LC3-positve compartments, whereas viable promastigotes are able to get rid of LC3-protein on their PV suggesting an involvement in parasite development and survival. In this thesis, we established a lentiviral based fluorescent imaging technique that we combined with High-Pressure-Freezing (HPF) and high-resolution 3D electron microscopy. We visualized a promastigote in a LC3-compartment whose ultrastructure showed an opening of the PV to the outside. To identify new LAP markers involved in Leishmania infection, we established an immuno-magnetic isolation protocol for the purification of Leishmania containing compartments.rnIn conclusion, this study suggests that L. major compartment biogenesis and maturation in pro- and anti-inflammatory human macrophages is dependent on the parasite stage and is different between axenic amastigotes, viable promastigotes and apoptotic promastigotes. Understanding the development and maturation of Leishmania parasites in human host cells is important to control and combat the neglected disease leishmaniasis in the future.rn

Biomimetic coating of organic polymers with a protein-functionalized layer of calcium phosphate: the surface properties of the carrier influence neither the coating characteristics nor the incorporation mechanism or release kinetics of the protein

Polymers that are used in clinical practice as bone-defect-filling materials possess many essential qualities, such as moldability, mechanical strength and biodegradability, but they are neither osteoconductive nor osteoinductive. Osteoconductivity can be conferred by coating the material with a layer of calcium phosphate, which can be rendered osteoinductive by functionalizing it with an osteogenic agent. We wished to ascertain whether the morphological and physicochemical characteristics of unfunctionalized and bovine-serum-albumin (BSA)-functionalized calcium-phosphate coatings were influenced by the surface properties of polymeric carriers. The release kinetics of the protein were also investigated. Two sponge-like materials (Helistat® and Polyactive®) and two fibrous ones (Ethisorb and poly[lactic-co-glycolic acid]) were tested. The coating characteristics were evaluated using state-of-the-art methodologies. The release kinetics of BSA were monitored spectrophotometrically. The characteristics of the amorphous and the crystalline phases of the coatings were not influenced by either the surface chemistry or the surface geometry of the underlying polymer. The mechanism whereby BSA was incorporated into the crystalline layer and the rate of release of the truly incorporated depot were likewise unaffected by the nature of the polymeric carrier. Our biomimetic coating technique could be applied to either spongy or fibrous bone-defect-filling organic polymers, with a view to rendering them osteoconductive and osteoinductive.

'Pergularain e I'--a plant cysteine protease with thrombin-like activity from Pergularia extensa latex

Pergularain e I, a cysteine protease with thrombin-like activity, was purified by ion exchange chromatography from the latex of Pergularia extensa. Its homogeneity was characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), native PAGE and reverse-phase high-performance liquid chromatography (RP-HPLC). The molecular mass of pergularain e I by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) was found to be 23.356 kDa and the N-terminal sequence is L-P-H-D-V-E. Pergularain e I is a glycoprotein containing approximately 20% of carbohydrate. Pergularain e I constituted 6.7% of the total protein with a specific activity of 9.5 units/mg/min with a 2.11-fold increased purity. Proteolytic activity of the pergularain e I was completely inhibited by iodoacetic acid (IAA). Pergularain e I exhibited procoagulant activity with citrated plasma and fibrinogen similar to thrombin. Pergularain e I increases the absorbance of fibrinogen solution in concentration-dependent and time-dependent manner. At 10 microg concentration, an absorbance of 0.48 was reached within 10 min of incubation time. Similar absorbance was observed when 0.2 NIH units of thrombin were used. Thrombin-like activity of pergularain e I is because of the selective hydrolysis of A alpha and B beta chains of fibrinogen and gamma-chain was observed to be insusceptible to hydrolysis. Molecular masses of the two peptide fragments released from fibrinogen due to the hydrolysis by pergularain e I at 5-min incubation time were found to be 1537.21 and 1553.29 and were in close agreement with the molecular masses of 16 amino acid sequence of fibrinopeptide A and 14 amino acid sequence of fibrinopeptide B, respectively. Prolonged fibrinogen-pergularain e I incubation releases additional peptides and their sequence comparison of molecular masses of the released peptides suggested that pergularain e I hydrolyzes specifically after arginine residues.

Rapid fusion and syncytium formation of heterologous cells upon expression of the FGFRL1 receptor

The fusion of mammalian cells into syncytia is a developmental process that is tightly restricted to a limited subset of cells. Besides gamete and placental trophoblast fusion, only macrophages and myogenic stem cells fuse into multinucleated syncytia. In contrast to viral cell fusion, which is mediated by fusogenic glycoproteins that actively merge membranes, mammalian cell fusion is poorly understood at the molecular level. A variety of mammalian transmembrane proteins, among them many of the immunoglobulin superfamily, have been implicated in cell-cell fusion, but none has been shown to actively fuse cells in vitro. Here we report that the FGFRL1 receptor, which is up-regulated during the differentiation of myoblasts into myotubes, fuses cultured cells into large, multinucleated syncytia. We used luciferase and GFP-based reporter assays to confirm cytoplasmic mixing and to identify the fusion inducing domain of FGFRL1. These assays revealed that Ig-like domain III and the transmembrane domain are both necessary and sufficient to rapidly fuse CHO cells into multinucleated syncytia comprising several hundred nuclei. Moreover, FGFRL1 also fused HEK293 and HeLa cells with untransfected CHO cells. Our data show that FGFRL1 is the first mammalian protein that is capable of inducing syncytium formation of heterologous cells in vitro.

Role of hypoxia and growth and differentiation factor-5 on differentiation of human mesenchymal stem cells towards intervertebral nucleus pulposus-like cells

There is evidence that mesenchymal stem cells (MSCs) can differentiate towards an intervertebral disc (IVD)-like phenotype. We compared the standard chondrogenic protocol using transforming growth factor beta-1 (TGFß) to the effects of hypoxia, growth and differentiation factor-5 (GDF5), and coculture with bovine nucleus pulposus cells (bNPC). The efficacy of molecules recently discovered as possible nucleus pulposus (NP) markers to differentiate between chondrogenic and IVD-like differentiation was evaluated. MSCs were isolated from human bone marrow and encapsulated in alginate beads. Beads were cultured in DMEM (control) supplemented with TGFß or GDF5 or under indirect coculture with bNPC. All groups were incubated at low (2 %) or normal (20 %) oxygen tension for 28 days. Hypoxia increased aggrecan and collagen II gene expression in all groups. The hypoxic GDF5 and TGFß groups demonstrated most increased aggrecan and collagen II mRNA levels and glycosaminoglycan accumulation. Collagen I and X were most up-regulated in the TGFß groups. From the NP markers, cytokeratin-19 was expressed to highest extent in the hypoxic GDF5 groups; lowest expression was observed in the TGFß group. Levels of forkhead box F1 were down-regulated by TGFß and up-regulated by coculture with bNPC. Carbonic anhydrase 12 was also down-regulated in the TGFß group and showed highest expression in the GDF5 group cocultured with bNPC under hypoxia. Trends in gene expression regulation were confirmed on the protein level using immunohistochemistry. We conclude that hypoxia and GDF5 may be suitable for directing MSCs towards the IVD-like phenotype.

Spindle cell oncocytoma of the pituitary gland with follicle-like component: organotypic differentiation to support its origin from folliculo-stellate cells

Spindle cell oncocytoma (SCO) is a rare, non-adenomatous tumor originating from the anterior pituitary gland. Composed of fusiform, mitochondrion-rich cells sharing several immunophenotypic and ultrastructural properties with folliculo-stellate cells (FSC), SCO has been proposed to represent a neoplastic counterpart of the latter. To date, however, SCO has failed to meet one criterion commonly used in histological-based taxonomy and diagnostics; that of recapitulating any of FSCs' morphologically defined developmental or physiological states. We describe a unique example of SCO wherein a conventional fascicular texture was seen coexisting with and organically merging into follicle-like arrangements. The sellar tumor of 2.7 × 2.6 × 2.5 cm was transphenoidally resected from a 55-year old female. Preoperative magnetic resonance imaging indicated an isointense, contrast enhancing mass with suprasellar extension. Histology showed multiple rudimentary to well-formed, follicle-like cavities on a classical spindle cell background; while all the participating cells exhibited an SCO immunophenotype, including positivity for S100 protein, vimentin, EMA, Bcl-2, and TTF-1, as well as staining with the antimitochondrial antibody 113-1. Conversely no expression of GFAP, follicular-epithelial cytokeratin, carcinoembryonic antigen, or anterior pituitary hormones was detected. Ultrastructurally, tumor cells facing follicular lumina displayed organelles of epithelial specialization, in particular surface microvilli and apical tight junctions. This constellation is felt to be reminiscent of FSCs' metaplastic transition to follicular epithelium, as observed during embryonic development and physiological renewal of the hormone-secreting parenchyma. Such finding is apt to being read as a supporting argument for SCO's descent from the FSC lineage.

Amicrobial pustulosis-like rash in a patient with Crohn's disease under anti-TNF-alpha blocker

Amicrobial pustulosis of the folds (APF) is a recently described entity characterized by relapsing pustular lesions predominantly involving the cutaneous flexures and scalp. This disease typically occurs in association with systemic lupus erythematosus and a variety of other autoimmune diseases. We here describe an APF-like pustular eruption predominantly affecting the scalp, face and trunk, occurring during long-term infliximab treatment for Crohn's disease. Immunohistochemical staining of skin biopsy specimens for myxovirus resistance protein A, a marker for type 1 interferon-inducible proteins, showed increased staining in the epidermis and dermal mononuclear inflammatory infiltrate. Our observation further extends the spectrum of cutaneous adverse reactions potentially related to anti-tumor necrosis factor-α, the clinical context in which APF can occur as well as its clinical presentations.

Ablation of the tumor suppressor histidine triad nucleotide binding protein 1 is protective against hepatic ischemia/reperfusion injury

The identification of cellular pathways capable of limiting ischemia/reperfusion (I/R) injury remains a frontier in medicine, and its clinical relevance is urgent. Histidine triad nucleotide binding protein 1 (HINT1) is a tumor suppressor that influences apoptosis. Because apoptotic pathways are a feature of I/R injury, we asked whether Hint1 influences hepatic I/R injury. Hint1(-/-) and C57BL/6 mice were subjected to 70% liver ischemia followed by reperfusion for 3 or 24 hours or to a sham operation. The serum aminotransferase levels, histological lesions, apoptosis, reactive oxygen species, and expression of B cell lymphoma 2-associated X protein (Bax), heme oxygenase 1 (HO-1), interleukin-6 (IL-6), IL-10, tumor necrosis factor-a, Src, nuclear factor kappa B (p65/RelA), and c-Jun were quantified. The responses to toll-like receptor ligands and nicotinamide adenine dinucleotide phosphate oxidase activity in Kupffer cells were compared in Hint1(-/-) mice and C57BL/6 mice. After I/R, the levels of serum aminotransferases, parenchymal necrosis, and hepatocellular apoptosis were significantly lower in Hint1(-/-) mice versus control mice. Furthermore, Bax expression decreased more than 2-fold in Hint1(-/-) mice, and the increases in reactive oxygen species and HO-1 expression that were evident in wild-type mice after I/R were absent in Hint1(-/-) mice. The phosphorylation of Src and the nuclear translocation of p65 were increased in Hint1(-/-) mice, whereas the nuclear expression of phosphorylated c-Jun was decreased. The levels of the protective cytokines IL-6 and IL-10 were increased in Hint1(-/-) mice. These effects increased survival after I/R in mice lacking Hint1. Hint1(-/-) Kupffer cells were less activated than control cells after stimulation with lipopolysaccharides. CONCLUSION: The Hint1 protein influences the course of I/R injury, and its ablation in Kupffer cells may limit the extent of the injury.

The formyl peptide receptor like-1 and scavenger receptor MARCO are involved in glial cell activation in bacterial meningitis

Recent studies have suggested that the scavenger receptor MARCO (macrophage receptor with collagenous structure) mediates activation of the immune response in bacterial infection of the central nervous system (CNS). The chemotactic G-protein-coupled receptor (GPCR) formyl-peptide-receptor like-1 (FPRL1) plays an essential role in the inflammatory responses of host defence mechanisms and neurodegenerative disorders such as Alzheimer's disease (AD). Expression of the antimicrobial peptide cathelicidin CRAMP/LL-37 is up-regulated in bacterial meningitis, but the mechanisms underlying CRAMP expression are far from clear.

Glycan-binding specificities of Streptococcus mutans and Streptococcus sobrinus lectin-like adhesins

Since the adhesion of bacteria to the tooth surface is a prerequisite for dental plaque and subsequent caries development, a promising caries preventive strategy could be to block the lectin-glycan-mediated adherence of cariogenic bacteria. The aim of the study was to evaluate potential differences in glycan-binding specificities of two Streptococcus mutans strains (DSM 20523 and DSM 6178) and Streptococcus sobrinus (DSM 20381). A competitive enzyme-linked lectin-binding assay was used to identify the binding specificities of isolated bacterial surface lectins. Blotting of the microbial proteins on neoglycoprotein-coated PVP membranes enabled a qualitative protein analysis of all specific bacterial lectins. Different glycan-binding sites could be identified for the S. mutans strains in comparison to S. sobrinus. An earlier reported glycan-binding specificity for terminal galactose residues could be confirmed for the S. mutans strains. For the S. sobrinus strain, more than one glycan-binding specificity could be found (oligomannose and terminal sialyl residues). Each of the tested strains showed more than one surface lectin responsible for the specific lectin-binding with varying molecular weight (S. mutans, 90/155 kDa and S. sobrinus, 35/45 kDa). The established experimental setup could be used as future standard procedure for the identification of bacterial lectin-derived binding specificities. The findings from this study might serve as basis for the design of an individual 'glycan cocktail' for the competitive inhibition of lectin-mediated adhesion of mutans streptococci to oral surfaces.

Besnoitia besnoiti protein disulfide isomerase (BbPDI): molecular characterization, expression and in silico modelling

Besnoitia besnoiti is an apicomplexan parasite responsible for bovine besnoitiosis, a disease with a high prevalence in tropical and subtropical regions and re-emerging in Europe. Despite the great economical losses associated with besnoitiosis, this disease has been underestimated and poorly studied, and neither an effective therapy nor an efficacious vaccine is available. Protein disulfide isomerase (PDI) is an essential enzyme for the acquisition of the correct three-dimensional structure of proteins. Current evidence suggests that in Neosporacaninum and Toxoplasmagondii, which are closely related to B. besnoiti, PDI play an important role in host cell invasion, is a relevant target for the host immune response, and represents a promising drug target and/or vaccine candidate. In this work, we present the nucleotide sequence of the B. besnoiti PDI gene. BbPDI belongs to the thioredoxin-like superfamily (cluster 00388) and is included in the PDI_a family (cluster defined cd02961) and the PDI_a_PDI_a'_c subfamily (cd02995). A 3D theoretical model was built by comparative homology using Swiss-Model server, using as a template the crystallographic deduced model of Tapasin-ERp57 (PDB code 3F8U chain C). Analysis of the phylogenetic tree for PDI within the phylum apicomplexa reinforces the close relationship among B. besnoiti, N. caninum and T. gondii. When subjected to a PDI-assay based on the polymerisation of reduced insulin, recombinant BbPDI expressed in E. coli exhibited enzymatic activity, which was inhibited by bacitracin. Antiserum directed against recombinant BbPDI reacted with PDI in Western blots and by immunofluorescence with B. besnoiti tachyzoites and bradyzoites.

Absence of the calcium-binding protein calretinin, not of calbindin D-28k, causes a permanent impairment of murine adult hippocampal neurogenesis

Calretinin (CR) and calbindin D-28k (CB) are cytosolic EF-hand Ca(2+)-binding proteins and function as Ca(2+) buffers affecting the spatiotemporal aspects of Ca(2+) transients and possibly also as Ca(2+) sensors modulating signaling cascades. In the adult hippocampal circuitry, CR and CB are expressed in specific principal neurons and subsets of interneurons. In addition, CR is transiently expressed within the neurogenic dentate gyrus (DG) niche. CR and CB expression during adult neurogenesis mark critical transition stages, onset of differentiation for CR, and the switch to adult-like connectivity for CB. Absence of either protein during these stages in null-mutant mice may have functional consequences and contribute to some aspects of the identified phenotypes. We report the impact of CR- and CB-deficiency on the proliferation and differentiation of progenitor cells within the subgranular zone (SGZ) neurogenic niche of the DG. Effects were evaluated (1) two and four weeks postnatally, during the transition period of the proliferative matrix to the adult state, and (2) in adult animals (3 months) to trace possible permanent changes in adult neurogenesis. The absence of CB from differentiated DG granule cells has no retrograde effect on the proliferative activity of progenitor cells, nor affects survival or migration/differentiation of newborn neurons in the adult DG including the SGZ. On the contrary, lack of CR from immature early postmitotic granule cells causes an early loss in proliferative capacity of the SGZ that is maintained into adult age, when it has a further impact on the migration/survival of newborn granule cells. The transient CR expression at the onset of adult neurogenesis differentiation may thus have two functions: (1) to serve as a self-maintenance signal for the pool of cells at the same stage of neurogenesis contributing to their survival/differentiation, and (2) it may contribute to retrograde signaling required for maintenance of the progenitor pool.