Journal: PLoS Genetics

Article Title: HOMER2, a Stereociliary Scaffolding Protein, Is Essential for Normal Hearing in Humans and Mice

doi: 10.1371/journal.pgen.1005137

Figure Lengend Snippet: (A) The pedigree of a five-generation family segregating progressive ADNSHL. DNA samples were obtained for 9 unaffected and 10 affected individuals. The partially filled symbol represents a two-year-old female who carries the HOMER2 mutation but in whom a formal ABR has not been performed. Individuals who underwent TGE+MPS using either the deafness panel (OtoSCOPE) or WES are marked with an O or W, respectively. Genotypes of participating family members are shown below each symbol in single letter amino acid nomenclature: P for Proline and R for Arginine. ( B) Age-related typical audiograms (ARTA). Binaural mean air conduction thresholds (dB HL) are presented for the ages 10–70 years. Hearing levels ranged from 0 to 120 dB depending on age and frequency; the annual threshold deterioration (ATD) was 1.2–1.6 dB/year at all frequencies. ( C) Representative chromatograms from wild-type and mutant sequences. ( D) Diagram of HOMER2 structure; the amino acid numbering indicates the beginning and end of the EVH1 and CC domains. The CC includes the CDC42 binding domain (CBD), Leucine Zipper-A (LZA) and Leucine Zipper-B (LZB). The position of the p.Arg185Pro mutation is shown in red.

Article Snippet: Following infiltration using 0.3% Triton X-100 and blocking with 5% normal goat serum, we incubated the tissues in rabbit HOMER2 polyclonal primary antibody (NB100-98712, Novus Biologicals, Littleton, CO) diluted 1:1000 in PBS overnight at 4°C.

Techniques: Mutagenesis, Binding Assay

Journal: PLoS Genetics

Article Title: HOMER2, a Stereociliary Scaffolding Protein, Is Essential for Normal Hearing in Humans and Mice

doi: 10.1371/journal.pgen.1005137

Figure Lengend Snippet: (A ) Staining with F-actin shows three rows of OHCs and one row of IHCs in the cochlea. ( B ) Homer2 staining in the OHCs and IHCs shows localization to stereocilia. ( C ) Merged pictures showing co-localization of Homer2 with F-actin in HC stereocilia. ( D ) Zoomed view of OHCs shows pronounced localization of Homer2 to the tips of stereocilia. Scale bar represents 10μm.

Article Snippet: Following infiltration using 0.3% Triton X-100 and blocking with 5% normal goat serum, we incubated the tissues in rabbit HOMER2 polyclonal primary antibody (NB100-98712, Novus Biologicals, Littleton, CO) diluted 1:1000 in PBS overnight at 4°C.

Techniques: Staining

Journal: PLoS Genetics

Article Title: HOMER2, a Stereociliary Scaffolding Protein, Is Essential for Normal Hearing in Humans and Mice

doi: 10.1371/journal.pgen.1005137

Figure Lengend Snippet: (A-D) Auditory brainstem responses (ABR) to broad band clicks (A) and tone bursts (B-D) at 8, 16, and 32 kHz in P14, P28 and P56 Homer2 -/- , Homer2 +/- and WT mice. (A) Raised mean ABR thresholds (dB SPL) were detected as early as P14 in Homer2 -/- mice and continued to increase with age. (B) At P14 Homer2 -/- mice had severe-to-profound hearing loss at high frequencies (16 kHz and 32 kHz) whereas hearing thresholds in their WT and Homer2 +/- littermates were in the normal range. (C) At P28 Homer2 -/- mice had hearing loss in the lower frequencies. (D) P56 Homer2 -/- mice had profound hearing loss across all tested frequencies (click, 8 kHz, 16 kHz and 32 kHz). (E-G) DPOAE levels. (E) In P14 Homer2 -/- mice, DPOAE amplitudes are significantly lower in the high frequencies (16, 22.6, and 32.0 kHz) as compared to their WT and Homer2 +/- littermates. (F) In P28 Homer2 -/- mice, DPOAE amplitudes are lower in nearly all frequencies (5.7, 8, 11.3, 16, 22.6, and 32.0 kHz). (G) In P56 Homer2 -/- mice, DPOAEs deteriorate across all frequencies consistent with profound hearing loss. (H) Representative Alexa-Fluor-488-phalloidin immunofluorescence shows no obvious hair cell death in cochleae in P56 Homer2 -/- (n = 5), Homer2 +/- (n = 4) and WT (n = 3) mice. Mean ABR thresholds and DPOAE amplitudes: Homer2 -/- mice are shown in red; Homer2 +/- in blue; and WT mice in green. The number of ears tested is shown in parentheses. P-values were calculated with One-way ANOVA and post-hoc T-test analysis. Asterisks indicate statistical significance: *P<0.05, **P<0.005, ***P<0.0005. Error bars represent SEM. Scale bar represents 10μm.

Article Snippet: Following infiltration using 0.3% Triton X-100 and blocking with 5% normal goat serum, we incubated the tissues in rabbit HOMER2 polyclonal primary antibody (NB100-98712, Novus Biologicals, Littleton, CO) diluted 1:1000 in PBS overnight at 4°C.

Techniques: Immunofluorescence

Journal: Pathology

Article Title: Overexpression of HOMER2 predicts better outcome in low-grade endometrioid endometrial adenocarcinoma

doi: 10.1016/j.pathol.2018.03.004

Figure Lengend Snippet: (A) Strong (2–3+) immunohistochemical staining with HOMER2. (B) Negative (0–1+) immunohistochemical staining with HOMER2.

Article Snippet: Antigen retrieval was performed by heating in the microwave for 20 min, before incubating the sections with a polyclonal recombinant protein HOMER2 antibody (1:50, 60 min; Novus Biologicals, USA).

Techniques: Immunohistochemical staining, Staining

Journal: Pathology

Article Title: Overexpression of HOMER2 predicts better outcome in low-grade endometrioid endometrial adenocarcinoma

doi: 10.1016/j.pathol.2018.03.004

Figure Lengend Snippet: Correlation of HOMER2 expression and clinical and pathological data

Article Snippet: Antigen retrieval was performed by heating in the microwave for 20 min, before incubating the sections with a polyclonal recombinant protein HOMER2 antibody (1:50, 60 min; Novus Biologicals, USA).

Techniques: Expressing

Journal: Pathology

Article Title: Overexpression of HOMER2 predicts better outcome in low-grade endometrioid endometrial adenocarcinoma

doi: 10.1016/j.pathol.2018.03.004

Figure Lengend Snippet: Survival curve for group 1 cohort showing better cause-specific survival in patients with a strong HOMER2 expression.

Article Snippet: Antigen retrieval was performed by heating in the microwave for 20 min, before incubating the sections with a polyclonal recombinant protein HOMER2 antibody (1:50, 60 min; Novus Biologicals, USA).

Techniques: Expressing

Journal: Pathology

Article Title: Overexpression of HOMER2 predicts better outcome in low-grade endometrioid endometrial adenocarcinoma

doi: 10.1016/j.pathol.2018.03.004

Figure Lengend Snippet: Multivariate analysis for endometrial cancer-specific survival in endometrioid adenocarcinoma

Article Snippet: Antigen retrieval was performed by heating in the microwave for 20 min, before incubating the sections with a polyclonal recombinant protein HOMER2 antibody (1:50, 60 min; Novus Biologicals, USA).

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: Distinct and Overlapping Expression Patterns of the Homer Family of Scaffolding Proteins and Their Encoding Genes in Developing Murine Cephalic Tissues

doi: 10.3390/ijms21041264

Figure Lengend Snippet: Expression patterns of Homer1, Homer2 and Homer3 during early stages of tooth formation. ( A – C ) Representative immunostaining of frontal sections across the first molars at embryonic day 12.5 (E12.5) showing the distribution of Homer1 ( A ), Homer2 ( B ) and Homer3 ( C ) proteins (purple). All three Homer proteins are expressed in the dental placode (epithelium) and Homer1 protein is concentrated in puncta. The jaw mesenchyme, including the mesenchyme adjacent to the dental placode, expresses Homer2 and Homer3 proteins and the vascular endothelium shows weak Homer1, moderate Homer2 and strong Homer3 immunostaining. ( D – O ) Representative in situ hybridization ( D – F , J – L ) and immunohistochemistry ( G – I , M – O ) data in frontal ( D – F , J – O ) and parasagittal ( G – I ) sections across the first molars at E13.5 ( D – I ; bud stage) and E14.5 ( J – O ; cap stage) showing expression of Homer1 , Homer2 and Homer3 transcripts (brown) and Homer1, Homer2 and Homer3 proteins (purple) in the dental epithelium and dental mesenchyme. Cells of the dental epithelium exhibit diffuse Homer immunostaining of the cytoplasm and membranes as well as puncta enriched with Homer proteins. The vascular endothelium is enriched with Homer3 protein and mRNA and exhibits weak Homer1 and moderate Homer2 immunostaining. The dotted line in ( D – F ) and ( J – L ) highlights the junction between the dental epithelium and the dental mesenchyme. BV, blood vessels; DE, dental epithelium; DPl, dental placode; DM, dental mesenchyme. Scale bars: 50 µm ( A – O ).

Article Snippet: The affinity-purified rabbit anti-Homer2 (product No. NBP1-85487) and the affinity-purified rabbit anti-Homer3 (product No. NBP2-32607) were from Novus Biologicals (Abingdon, UK).

Techniques: Expressing, Immunostaining, In Situ Hybridization, Immunohistochemistry

Journal: International Journal of Molecular Sciences

Article Title: Distinct and Overlapping Expression Patterns of the Homer Family of Scaffolding Proteins and Their Encoding Genes in Developing Murine Cephalic Tissues

doi: 10.3390/ijms21041264

Figure Lengend Snippet: Expression patterns of Homer1, Homer2 and Homer3 during the bell stage of tooth development. ( A – F ) Representative frontal ( A ) and parasagittal ( B – F ) sections across developing first molars at 1 day postpartum (1 dpp) after in situ hybridization ( A – C ) and immunohistochemistry ( D – F ) for Homer1 ( A , D ), Homer2 ( B , E ) and Homer3 ( C , F ). ( A’ – F’ ) are magnified views of the boxed areas in ( A – F ). Odontoblasts and differentiating ameloblasts are enriched with Homer proteins (purple) and mRNAs (brown) as compared to the dental papilla, stellate reticulum and inner dental epithelium. Homer2 protein and mRNA are also expressed in the stratum intermedium. Homer proteins are concentrated in intracellular puncta in odontoblasts and differentiating ameloblasts as well as in cells of the stratum intermedium and inner dental epithelium. The endothelium of blood vessels in the stellate reticulum and dental mesenchyme is enriched with Homer3 mRNA and protein and exhibits weak and moderate immunostaining for Homer1 and Homer2, respectively. Homer proteins are also detectable in osteoblasts within the developing alveolar bone ( D – F ). BV, blood vessels; dAM, differentiating ameloblasts; DP, dental papilla; IDE, inner dental epithelium; Ob, osteoblasts; Od, odontoblasts; pOd, preodontoblasts; SI, stratum intermedium; SR, stellate reticulum. Scale bars: 200 µm ( A – F ) and 100 µm ( A’ – F’ ).

Article Snippet: The affinity-purified rabbit anti-Homer2 (product No. NBP1-85487) and the affinity-purified rabbit anti-Homer3 (product No. NBP2-32607) were from Novus Biologicals (Abingdon, UK).

Techniques: Expressing, In Situ Hybridization, Immunohistochemistry, Immunostaining

Journal: International Journal of Molecular Sciences

Article Title: Distinct and Overlapping Expression Patterns of the Homer Family of Scaffolding Proteins and Their Encoding Genes in Developing Murine Cephalic Tissues

doi: 10.3390/ijms21041264

Figure Lengend Snippet: Expression patterns of Homer1, Homer2 and Homer3 during advanced stages of tooth formation. ( A – R ) Representative in situ hybridization ( A – C , G – I ) and immunohistochemistry ( D – F , J – R ) data showing the distribution patterns of Homer1 ( A , G ), Homer2 ( B , H ) and Homer3 ( C , I ) transcripts (brown) and Homer1 ( D , J , M , P ), Homer2 ( E , K , N , Q ) and Homer3 ( F , L , O , R ) proteins (purple) in developing teeth at 12 days postpartum (12 dpp). ( A – F ) Sections across incisors at the level of the secretory stage of enamel formation. Secretory ameloblasts and young odontoblasts (cells facing a thin layer of predentin/dentin matrices) express the three Homer proteins and their encoding genes. The asterisks in ( D , F ) mark an artefactual space due to detachment of secretory ameloblasts from dentin. ( G – L ) Sections at the level of enamel maturation showing expression of the three Homer proteins and their encoding genes in maturation-stage ameloblasts and in the papillary layer. ( M – R ) Sections across molars showing expression of Homer proteins in mature odontoblasts (cells that have produced a thick layer of predentin/dentin) ( M – O ) and in Hertwig’s epithelial root sheath ( P – R ). Secretory ameloblasts, maturation-stage ameloblasts, odontoblasts and cells of the papillary layer exhibit puncta enriched with Homer proteins ( D – F , J – O ). Note that the Homer-positive(+) puncta in young odontoblasts ( D – F ) are less prominent than the Homer+ puncta in mature odontoblasts ( M – O ) and that the Homer3+ puncta in subsets of cells of the dental pulp are apparently smaller than the Homer3+ puncta in mature odontoblasts ( O ). Odontoblasts in the developing roots express Homer proteins ( P – R ). The endothelium of vascular loops penetrating the papillary layer ( I , L ) and of blood vessels in the dental pulp and dental sac mesenchyme ( C , F , I , L , O , R ) expresses Homer3 mRNA and protein and exhibits weak Homer1 ( D , J , M , P ) and moderate Homer2 ( E , K , N , Q ) immunostaining. BV, blood vessels; DS, dental sac; HS, Hertwig’s epithelial root sheath; MA, maturation-stage ameloblasts; Od, odontoblasts; P, dental pulp; PD/D, predentin/dentin matrices; PL, papillary layer; SA, secretory ameloblasts; SI, stratum intermedium. Scale bars: 50 µm ( A – R ).

Article Snippet: The affinity-purified rabbit anti-Homer2 (product No. NBP1-85487) and the affinity-purified rabbit anti-Homer3 (product No. NBP2-32607) were from Novus Biologicals (Abingdon, UK).

Techniques: Expressing, In Situ Hybridization, Immunohistochemistry, Produced, Immunostaining

Journal: International Journal of Molecular Sciences

Article Title: Distinct and Overlapping Expression Patterns of the Homer Family of Scaffolding Proteins and Their Encoding Genes in Developing Murine Cephalic Tissues

doi: 10.3390/ijms21041264

Figure Lengend Snippet: Expression patterns of Homer1, Homer2 and Homer3 in the developing forebrain and trigeminal ganglion. ( A – I ) Representative sections across the forebrain and trigeminal ganglion at embryonic day 14.5 (E14.5) after in situ hybridization revealing Homer transcripts ( A – C ) and immunostaining ( D – I ) for Homer proteins. Homer transcripts (brown) and Homer proteins (purple) are expressed in the trigeminal ganglion and in different regions of the forebrain, including the striatum, neocortex, thalamus, hypothalamus and in the hippocampal formation, and Homer proteins are enriched in differentiating fields and in the apical surface of the ventricular layer. ( G’ – I’ ) are magnified views of boxed areas in ( G – I ) showing that Homer1, Homer2 and Homer3 proteins are enriched in the apical surface of cells of the choroid plexus and that in these cells Homer1 and Homer3 are also concentrated in puncta. ( G’’ – I’’ ) are magnified views of boxed areas in ( G – I ) showing the distribution of Homer proteins in the anterior hypothalamic area and concentration of Homer1 and Homer2 in intracellular puncta. The three Homer proteins are also detectable in the apical surface of the hypothalamic neuroepithelium ( G’’ – H’’ ). ( G’’’ – I’’’ ) are magnified views of boxed areas in ( G – I ), showing that in other brain regions, the intracellular Homer-positive(+) puncta are less conspicuous than the Homer+ puncta in the anterior hypothalamic area. The endothelial lining of blood vessels in the brain and meninges expresses Homer3 transcripts and proteins ( C , F , I – I’’’ ) and exhibits weak Homer1 ( D , G – G’’’ ) and moderate Homer2 immunostaining ( E , H – H’’’ ). BV, blood vessels; Ch, choroid plexus; Cx, neocortex; Hc, hippocampal formation; Ht, hypothalamus; Hn, hypothalamic neuroepithelium; Lv, lateral ventricle; St, striatum; Th, thalamus; Vg, trigeminal ganglion; V3, third ventricle. Scale bars: 500 µm ( A – I ) and 50 µm ( G’ – I’’’ ).

Article Snippet: The affinity-purified rabbit anti-Homer2 (product No. NBP1-85487) and the affinity-purified rabbit anti-Homer3 (product No. NBP2-32607) were from Novus Biologicals (Abingdon, UK).

Techniques: Expressing, In Situ Hybridization, Immunostaining, Concentration Assay

Journal: International Journal of Molecular Sciences

Article Title: Distinct and Overlapping Expression Patterns of the Homer Family of Scaffolding Proteins and Their Encoding Genes in Developing Murine Cephalic Tissues

doi: 10.3390/ijms21041264

Figure Lengend Snippet: Expression patterns of Homer1, Homer2 and Homer3 in neuronal and non-neuronal cephalic tissues. ( A – I ) Representative sections across cephalic tissues at embryonic day 14.5 (E14.5) after in situ hybridization ( A – C ) and immunohistochemistry ( D – I ). ( A’ – C’ ) and ( D’ – F’ ) are magnified views of the boxed areas in ( A – C ) and ( D – F ), respectively. ( A – F’ ) The Homer1 , Homer2 and Homer3 mRNA expression patterns (brown) are consistent with the distribution patterns of their protein products (purple) in several cephalic structures, including the olfactory bulbs, retina, lens epithelium, olfactory epithelium, submandibular salivary glands, tooth, rugae palatinae, skeletal muscle and medial epithelial seam of the secondary palate. Note that Homer2 mRNA is enriched in developing muscles of the tongue (arrowheads in B’ ). The olfactory epithelium displays strong Homer2 ( B , E ) but moderate Homer1 ( A , D ) and Homer3 ( C , F ) hybridization signals and immunolabelling. The olfactory and trigeminal nerves show moderate Homer1 ( D , D’ ) and strong Homer2 ( E , E’ ) immunostaining. Homer3 immunoreactivity is strong in olfactory nerves and moderate in trigeminal nerves ( F , F’ ). ( G – I ) Homer1, Homer2 and Homer3 proteins are enriched in intracellular puncta in the epithelium of developing submandibular glands and the three Homer proteins are detectable in nerves within the glandular stroma. Note that vascular endothelium in cephalic tissues expresses Homer3 mRNA and protein and exhibits weak Homer1 and moderate Homer2 immunostaining. ( J – L ) Representative sections across submandibular salivary glands at 1 day postpartum (1 dpp) after immunostaining for Homer1 ( J ), Homer2 ( K ) and Homer3 ( L ). ( J’ – L’ ) are magnified views of the boxed areas in ( J – K ). In epithelial cells (acinar and tubular cells) of the glands Homer proteins show overlapping and distinct subcellular distribution patterns. Homer1 is enriched in the apical surface/membranes and in numerous intracellular puncta ( J , J’ ); Homer2 is concentrated in the apical surface/membranes and is also detectable in the cytoplasm ( K , K’ ); and Homer3 is detectable in the cytoplasm, cell membranes and puncta ( L , L’ ). The Homer2-positive(+) and Homer3+ puncta are less conspicuous than the Homer1+ puncta in glandular epithelial cells ( J – L’ ) and the Homer+ puncta in 1 dpp glands ( J – K ) are relatively smaller than the Homer+ puncta in E14.5 glands ( G – I ). BV, blood vessels; L, lens epithelium; MES, medial epithelial seam; Nf, nerve fibers (in salivary glands); n5, trigeminal nerve; OB, olfactory bulb; OE, olfactory epithelium; ON, olfactory nerve; R, retina; RP, rugae palatinae; SG, submandibular salivary glands; SM, skeletal muscle (masseter); T, tongue; To, tooth. Scale bars: 1mm ( A – F ), 200 µm ( A’ – F’ ), 50 µm ( G – L ) and 10 µm ( J’ – L’ ).

Article Snippet: The affinity-purified rabbit anti-Homer2 (product No. NBP1-85487) and the affinity-purified rabbit anti-Homer3 (product No. NBP2-32607) were from Novus Biologicals (Abingdon, UK).

Techniques: Expressing, In Situ Hybridization, Immunohistochemistry, Muscles, Hybridization, Immunostaining

Journal: International Journal of Molecular Sciences

Article Title: Distinct and Overlapping Expression Patterns of the Homer Family of Scaffolding Proteins and Their Encoding Genes in Developing Murine Cephalic Tissues

doi: 10.3390/ijms21041264

Figure Lengend Snippet: Expression patterns of Homer1, Homer2 and Homer3 in the olfactory and respiratory mucosae and in the cochlea. ( A – O ) Representative immunohistochemistry ( A – F , J – O ) and in situ hybridization ( G – I ) data in sections across the nasal cavity ( A – F ) and cochlea ( G – O ) at 1 day postpartum (1 dpp) showing the distribution patterns of Homer proteins (purple) and their encoding genes (brown). ( A – C ) Homer1 ( A ), Homer2 ( B ) and Homer3 ( C ) proteins are enriched in the apical surface (likely cilia of olfactory neurons and microvilli of supporting cells) of cells of the olfactory epithelium and are also detectable in intracellular puncta. Dendrites (arrowheads in A – C ) and axons, the latter forming the olfactory nerves, are Homer1-positive(+), Homer2+ and Homer3+, indicating that some of the Homer+ cells in the olfactory epithelium are olfactory neurons. ( D – F ) The respiratory epithelium and nasal glands express Homer1 ( D ), Homer2 ( E ) and Homer3 ( F ) proteins. In cells of the respiratory epithelium the Homer1+ puncta are more prominent than the Homer2+ and Homer3+ puncta. In nasal gland acinar cells Homer1 is enriched in the apical membrane and in puncta, Homer2 is readily detectable in cell membranes and in the cytoplasm and Homer3+ puncta are detectable in subsets of acinar cells. Note that the Homer1+ puncta in nasal gland acinar cells are small compared to the Homer1+ puncta in cells of the respiratory epithelium ( D ). The endothelial lining of blood vessels in the olfactory ( A – C ) and respiratory ( D – F ) mucosae shows weak Homer1, moderate Homer2 and strong Homer3 immunolabelling. ( G – O ) Expression patterns of Homer transcripts ( G – I ) and their protein products ( J – O ) in the cochlea. Homer1 ( G ), Homer2 ( H ) and Homer3 ( I ) mRNAs are detectable in various cells of the developing organ of Corti, including inner (IHC) and outer (OHC) hair cells as well as in cells forming the stria vascularis, basilar membrane and Reissner’s membrane ( G – I ). Homer1 and Homer2 transcripts are enriched in IHC and OHC ( G , H ). Homer1 ( J ), Homer2 ( K ) and Homer3 ( L ) proteins show overlapping but also distinct subcellular localization in cells of the organ of Corti. In IHC and OHC Homer1 and Homer2 are enriched in the apical surface (likely in stereocilia) and in perinuclear puncta and are also detectable in the cytoplasm. By contrast, Homer3 protein is detectable in puncta at the apical surface of IHC and OHC and in perinuclear puncta in IHC. Other cells of the organ of Corti, including cells of the greater epithelial ridge (GER), cells of Claudius and Deiter’s cells, exhibit Homer1+ and Homer3+ perinuclear puncta and show moderate Homer1, very weak Homer2 and weak Homer3 immunostaining in the cytoplasm. Homer1 is also enriched in the apical (endolymphatic) surface of the GER. Cells of the stria vascularis and Reissner’s membrane exhibit Homer1+ and Homer3+ puncta and cytoplasmic Homer2 immunostaining. Note that in IHC and OHC the Homer1+ puncta are large as compared to the Homer2+ and Homer3+ puncta and that the Homer1+ puncta in IHC and OHC are large compared to the Homer1+ puncta in the cells of Claudius and in cells of the GER and stria vascularis. ( M – O ) Neurons of the cochlear spiral ganglion show diffuse cytoplasmic immunostaining for Homer1 ( M ), Homer2 ( N ) and Homer3 ( O ) and are enriched with Homer1+ and Homer3+ puncta. Note that the vascular endothelium in the cochlea exhibits weak Homer1, moderate Homer2 and strong Homer3 immunolabelling ( J – O ). BM, basilar membrane; BV, blood vessels; Cc, cells of Claudius; Dc, Deiter’s cells; GER, greater epithelial ridge; IHC, inner hair cells; NG, nasal glands; OE, olfactory epithelium; ON, olfactory nerve; RE, respiratory epithelium; RM, Reissner’s membrane; Sg, spiral ganglion; SV, stria vascularis. Scale bars: 100 µm ( A – C ) and 50 µm ( D – O ).

Article Snippet: The affinity-purified rabbit anti-Homer2 (product No. NBP1-85487) and the affinity-purified rabbit anti-Homer3 (product No. NBP2-32607) were from Novus Biologicals (Abingdon, UK).

Techniques: Expressing, Immunohistochemistry, In Situ Hybridization, Membrane, Immunostaining

Journal: International Journal of Molecular Sciences

Article Title: Distinct and Overlapping Expression Patterns of the Homer Family of Scaffolding Proteins and Their Encoding Genes in Developing Murine Cephalic Tissues

doi: 10.3390/ijms21041264

Figure Lengend Snippet: Expression patterns of Homer1, Homer2 and Homer3 in the tongue and alveolar bone. ( A – O ) Representative in situ hybridization ( A – C , G – I ) and immunohistochemistry ( D – F , J – O ) data showing the distribution patterns of Homer transcripts (brown) and Homer proteins (purple). ( A – F ) Sections across the developing tongue at embryonic day 14.5 (E14.5). Homer1 mRNA ( A ) and Homer1 protein ( D ) are enriched in developing taste buds and are detectable in the lingual epithelium (LE), developing muscles of the tongue (arrows) and in the lingual mesenchyme (LM). Homer2 mRNA ( B ) and Homer2 protein ( E ) are expressed in the LE, including in developing taste buds and are enriched in developing muscle of the tongue (arrows). Homer2 protein is also detectable in nerves innervating developing fungiform papillae (arrowhead in E ). Homer3 mRNA ( C ) and Homer3 protein ( F ) are expressed in the LE, LM and in the vascular endothelium, whereas developing taste buds are virtually devoid of Homer3 immunostaining. ( G – L ) Sections across the tongue at 1 day postpartum (1 dpp). Homer1 mRNA ( G ) and Homer1 protein ( J ) are enriched in nerves innervating fungiform papillae (arrowheads in G , J ) and in taste buds. The LE and taste buds express Homer2 mRNA ( H ) and Homer2 protein ( K ) and exhibit weak hybridization signals ( I ) and immunostaining (L ) for Homer3. By contrast, Homer3 transcripts and Homer3 protein are readily detectable in the vascular endothelium. ( M – O ) sections across the alveolar bone at 12 dpp. Homer1 ( M ), Homer2 ( N ) and Homer3 ( O ) proteins are expressed in osteoblasts (arrows in M , N , O ) and osteoclasts and Homer2 and Homer3 but not Homer1, are also detectable in osteocytes (arrowheads in M , N , O ). Note that in bone cells Homer proteins are enriched in puncta. BV, blood vessels; LE, lingual epithelium; LM, lingual mesenchyme; OC, osteoclast; TB, taste bud; SM, muscles of the tongue. Scale bars: 50 µm ( A – O ).

Article Snippet: The affinity-purified rabbit anti-Homer2 (product No. NBP1-85487) and the affinity-purified rabbit anti-Homer3 (product No. NBP2-32607) were from Novus Biologicals (Abingdon, UK).

Techniques: Expressing, In Situ Hybridization, Immunohistochemistry, Muscles, Immunostaining, Hybridization

Journal: Nature neuroscience

Article Title: Synaptic depression via mGluR1 positive allosteric modulation suppresses cue-induced cocaine craving

doi: 10.1038/nn.3590

Figure Lengend Snippet: ( a ) Timeline. SA, self-administration. ( b ) Training data for Saline and Cocaine groups (see legend to for description of training). ( c ) Cell surface and total protein levels of mGluR1 and mGluR5 were measured on three withdrawal days (WD): before (WD14), at the onset (WD25) and after (WD48) elevation of CP-AMPAR levels. Data are expressed as percent of Saline group (± s.e.m.) at each time-point (n values are provided within each bar). *p=0.04 (surface mGluR1, WD25), *p=0.03 (surface mGluR1, WD48), *p=0.03 (total mGluR1, WD48; t 18 =2.0), and *p=0.03 (surface mGluR5, WD48) versus respective saline groups. Differences in optical density for Saline groups on different withdrawal days reflect differences in exposure time for different blots rather than time-dependent changes in protein levels after saline self-administration. Full-length blots are presented in .

Article Snippet: For immunoblotting after IP, the same primary antibodies were used with two exceptions: mGluR1a (1:3,000; 2031-mGluR1a, PhosphoSolutions) and Homer2 (1:350, H00009455-B01P, Abnova).

Techniques: Saline

Journal: Nature neuroscience

Article Title: Synaptic depression via mGluR1 positive allosteric modulation suppresses cue-induced cocaine craving

doi: 10.1038/nn.3590

Figure Lengend Snippet: (a) Co-immunoprecipitation (co-IP) experiments assessing the physical associations between mGluR1 and Homer proteins on withdrawal day (WD) 14, 25 or 48 from extended-access cocaine (coc) or saline (sal) self-administration (see timeline in ). No significant changes in association between mGluR1 and Homer proteins were observed at any of the 3 withdrawal time-points. ( b ) In the case of mGluR5, no change in association with Homer proteins was observed at the two earlier withdrawal time-points (WD14 and WD25). However, a significant decrease in association between mGluR5 and both Homer isoforms was found on WD48 in animals that previously self-administered cocaine. Data are expressed as percent of Saline group (± s.e.m.) at each time-point (n values are provided within each bar). **p=0.01 (Homer1bc) and *p=0.04 (Homer2) versus respective Saline groups. Full-length blots are presented in .

Article Snippet: For immunoblotting after IP, the same primary antibodies were used with two exceptions: mGluR1a (1:3,000; 2031-mGluR1a, PhosphoSolutions) and Homer2 (1:350, H00009455-B01P, Abnova).

Techniques: Immunoprecipitation, Co-Immunoprecipitation Assay, Saline

Journal: Nature neuroscience

Article Title: Synaptic depression via mGluR1 positive allosteric modulation suppresses cue-induced cocaine craving

doi: 10.1038/nn.3590

Figure Lengend Snippet: ( a ) Timeline (SA, self-administration). Following completion of cocaine self-administration training, rats received daily injections with vehicle or an mGluR1 antagonist (JNJ16259685; 5 mg/kg, i.p.) on WD10-15 (2 rats) or WD11-16 (1 rat) and patch-clamp recordings were performed 1 day after the last injection (WD16 or WD17). ( b ) Left: In rats treated with vehicle during withdrawal, the RI on WD16-17 is similar to saline controls, whereas it is elevated after repeated JNJ injections. Each circle represents one cell. Lines are means for each group. Right: Representative traces of EPSC -70mV , EPSC +40mV and the reversal potential (in gray, 0 mV) for cells from Cocaine/Vehicle (Vehicle) and Cocaine/JNJ (JNJ) groups (calibration bar=50 pA, 40 ms). Overall, the reversal potentials did not differ significantly between the groups (p=0.18). ( c ) Repeated JNJ injections also increased naspm sensitivity on WD16-17 compared to the Coc/Vehicle group. Data are shown as group average EPSC -70mV amplitude (± s.e.m.) before (5 min) and during (15 min) Naspm application (left) and as the average percentage change from baseline during the last 5 min of Naspm application (right; each circle represents one cell; lines are means for each group) ***p<0.001, **p=0.002 versus Vehicle; Vehicle, n=8 cells/3 rats; JNJ, n=7 cells/3 rats.

Article Snippet: For immunoblotting after IP, the same primary antibodies were used with two exceptions: mGluR1a (1:3,000; 2031-mGluR1a, PhosphoSolutions) and Homer2 (1:350, H00009455-B01P, Abnova).

Techniques: Patch Clamp, Injection, Saline