A novel gene trap line for visualization and manipulation of erbb3b+ neural crest and glial cells in zebrafish.

Glia are a diverse and essential cell type in the vertebrate nervous system. Transgenic tools and fluorescent reporter lines are critical resources to investigate how glial subtypes develop and function. However, despite the many lines available in zebrafish, the community still lacks the ability to label all unique stages of glial development and specific subpopulations of cells. To address this issue, we screened zebrafish gene and enhancer trap lines to find a novel reporter for peripheral glial subtypes. From these, we generated the gSAIzGFFD37A transgenic line that expresses GFP in neural crest cells and central and peripheral glia. We found that the gene trap construct is located within an intron of erbb3b, a gene essential for glial development. Additionally, we confirmed that GFP+ â€‹cells express erbb3b along with sox10, a known glial marker. From our screen, we have identified the gSAIzGFFD37A line as a novel and powerful tool for studying glia in the developing zebrafish, as well as a new resource to manipulate erbb3b+ â€‹cells.

Sox10-cre BAC transgenes reveal temporal restriction of mesenchymal cranial neural crest and identify glandular Sox10 expression.

Diversity of neural crest derivatives has been studied with a variety of approaches during embryonic development. In mammals Cre-LoxP lineage tracing is a robust means to fate map neural crest relying on cre driven from regulatory elements of early neural crest genes. Sox10 is an essential transcription factor for normal neural crest development. A variety of efforts have been made to label neural crest derivatives using partial Sox10 regulatory elements to drive cre expression. To date published Sox10-cre lines have focused primarily on lineage tracing in specific tissues or during early fetal development. We describe two new Sox10-cre BAC transgenes, constitutive (cre) and inducible (cre/ERT2), that contain the complete repertoire of Sox10 regulatory elements. We present a thorough expression profile of each, identifying a few novel sites of Sox10 expression not captured by other neural crest cre drivers. Comparative mapping of expression patterns between the Sox10-cre and Sox10-cre/ERT2 transgenes identified a narrow temporal window in which Sox10 expression is present in mesenchymal derivatives prior to becoming restricted to neural elements during embryogenesis. In more caudal structures, such as the intestine and lower urinary tract, our Sox10-cre BAC transgene appears to be more efficient in labeling neural crest-derived cell types than Wnt1-cre. The analysis reveals consistent expression of Sox10 in non-neural crest derived glandular epithelium, including salivary, mammary, and urethral glands of adult mice. These Sox10-cre and Sox10-cre/ERT2 transgenic lines are verified tools that will enable refined temporal and cell-type specific lineage analysis of neural crest derivatives as well as glandular tissues that rely on Sox10 for proper development and function.

Adding Perplexity to Rarity: Diffuse S100-Protein and SOX10 Expression in a Molecularly Confirmed PAX7-Positive Primary Cutaneous Ewing Sarcoma.

ABSTRACT: Primary cutaneous Ewing sarcoma (EWS) is a very rare neoplasm that shares similar morphologic, immunohistochemical, and molecular features with its osseous counterpart. Herein, we present an extraordinarily rare case of PAX7-positive cutaneous EWS in a 9-year-old girl that was also diffusely positive for SOX10 and S100-protein. Next generation sequencing detected the EWSR1-FLI1 fusion supporting the diagnosis, which was further validated by break-apart EWSR1 fluorescence in situ hybridization. Diffuse S100-protein and SOX10 expression has been reported only in a handful of cases of EWS and may pose significant diagnostic challenges for dermatopathologists. PAX7 is a recently introduced marker, which is highly sensitive for EWS and can potentially have discriminatory power in the differential diagnosis of cutaneous undifferentiated round blue cell tumors.

A novel group of spindle cell tumors defined by S100 and CD34 co-expression shows recurrent fusions involving RAF1, BRAF, and NTRK1/2 genes.

Tumors characterized by co-expression of S100 and CD34, in the absence of SOX10, remain difficult to classify. Triggered by a few index cases with monomorphic cytomorphology and distinctive stromal and perivascular hyalinization, immunopositivity for S100 and CD34, and RAF1 and NTRK1 fusions, the authors undertook a systematic review of tumors with similar features. Most of the cases selected were previously diagnosed as low-grade malignant peripheral nerve sheath tumors, while others were deemed unclassified. The tumors were studied with targeted RNA sequencing and/or FISH. A total of 25 cases (15 adults and 10 children) with kinase fusions were identified, including 8 cases involving RAF1, 2 BRAF, 14 NTRK1, and 1 NTRK2 gene rearrangements. Most tumors showed a monomorphic spindle cell proliferation with stromal and perivascular keloidal collagen, in a patternless architecture, with only occasional scattered pleomorphic or multinucleated cells. Most cases showed low cellularity, a low mitotic count, and absence of necrosis. Although a subset showed overlap with lipofibromatosis-like neural tumors, the study group showed distinctive hyalinization and overt malignant features, such as highly cellular fascicular growth and primitive appearance. All tumors showed co-expression of S100 and CD34, ranging from focal to diffuse. SOX10 was negative in all cases. NTRK1 immunohistochemistry showed high levels of expression in all tumors with NTRK1 gene rearrangements. H3K27me3 expression performed in a subset of cases was retained. These findings together with the recurrent gene fusions in RAF1, BRAF, and NTRK1/2 kinases suggest a distinct molecular tumor subtype with consistent S100 and CD34 immunoreactivity.

Sox8 and Sox10 jointly maintain myelin gene expression in oligodendrocytes.

In Schwann cells of the vertebrate peripheral nervous system, induction of myelination and myelin maintenance both depend on the HMG-domain-containing transcription factor Sox10. In oligodendrocytes of the central nervous system, Sox10 is also essential for the induction of myelination. Its role in late phases of myelination and myelin maintenance has not been studied so far. Here, we show that these processes are largely unaffected in mice that lack Sox10 in mature oligodendrocytes. As Sox10 is co-expressed with the related Sox8, we also analyzed oligodendrocytes and myelination in Sox8-deficient mice. Again, we could not detect any major abnormalities. Expression of many myelin genes was only modestly reduced in both mouse mutants. Dramatic reductions in expression levels and phenotypic disturbances became only apparent once Sox8 and Sox10 were both absent. This argues that Sox8 and Sox10 are jointly required for myelin maintenance and impact myelin gene expression. One direct target gene of both Sox proteins is the late myelin gene Mog. Our results point to at least partial functional redundancy between both related Sox proteins in mature oligodendrocytes and are the first report of a substantial function of Sox8 in the oligodendroglial lineage.

A quantitative comparison between SOX10 and MART-1 immunostaining to detect melanocytic hyperplasia in chronically sun-damaged skin.

Histologic differentiation of melanoma in situ (MIS) from solar keratosis on chronically sun-damaged skin is challenging. The first-line immunostain is usually MART-1/Melan-A, which can exaggerate the epidermal melanocytes, causing a diagnostic pitfall for MIS. By comparing MART-1 and SOX10 immunostaining, we scored the percentage of epidermal melanocytes per 2-mm diameter fields in pigmented actinic keratosis (n = 16), lichenoid keratosis (n = 7), junctional melanocytic nevus (n = 6), keratosis with atypical melanocytic proliferation (n = 17) and MIS (n = 10). These cases represented an older population (68 years median age) and the head and neck (50%) was the most common anatomic site. MART-1 score was significantly higher than SOX10 (P value <.05) in solar keratoses, but showed no difference in detecting melanocytic proliferations, demonstrating their equal detection rate of melanocytes. The sensitivity of both MART-1 and SOX10 was 100%, while their specificities were 17% and 96%, respectively. These results show that SOX10 is more specific than MART-1 in distinguishing epidermal melanocytes on sun-damaged skin by avoiding overdiagnosis of melanoma.

Antagonistic cross-regulation between Sox9 and Sox10 controls an anti-tumorigenic program in melanoma.

Melanoma is the most fatal skin cancer, but the etiology of this devastating disease is still poorly understood. Recently, the transcription factor Sox10 has been shown to promote both melanoma initiation and progression. Reducing SOX10 expression levels in human melanoma cells and in a genetic melanoma mouse model, efficiently abolishes tumorigenesis by inducing cell cycle exit and apoptosis. Here, we show that this anti-tumorigenic effect functionally involves SOX9, a factor related to SOX10 and upregulated in melanoma cells upon loss of SOX10. Unlike SOX10, SOX9 is not required for normal melanocyte stem cell function, the formation of hyperplastic lesions, and melanoma initiation. To the contrary, SOX9 overexpression results in cell cycle arrest, apoptosis, and a gene expression profile shared by melanoma cells with reduced SOX10 expression. Moreover, SOX9 binds to the SOX10 promoter and induces downregulation of SOX10 expression, revealing a feedback loop reinforcing the SOX10 low/SOX9 high ant,m/ii-tumorigenic program. Finally, SOX9 is required in vitro and in vivo for the anti-tumorigenic effect achieved by reducing SOX10 expression. Thus, SOX10 and SOX9 are functionally antagonistic regulators of melanoma development.

The positive transcriptional elongation factor (P-TEFb) is required for neural crest specification.

Regulation of gene expression at the level of transcriptional elongation has been shown to be important in stem cells and tumour cells, but its role in the whole animal is only now being fully explored. Neural crest cells (NCCs) are a multipotent population of cells that migrate during early development from the dorsal neural tube throughout the embryo where they differentiate into a variety of cell types including pigment cells, cranio-facial skeleton and sensory neurons. Specification of NCCs is both spatially and temporally regulated during embryonic development. Here we show that components of the transcriptional elongation regulatory machinery, CDK9 and CYCLINT1 of the P-TEFb complex, are required to regulate neural crest specification. In particular, we show that expression of the proto-oncogene c-Myc and c-Myc responsive genes are affected. Our data suggest that P-TEFb is crucial to drive expression of c-Myc, which acts as a 'gate-keeper' for the correct temporal and spatial development of the neural crest.

MYB, CD117 and SOX-10 expression in cutaneous adnexal tumors.

BACKGROUND: Elevated MYB expression has been documented in adenoid cystic carcinoma (ACC), cylindroma, and spiradenoma, but the specificity of this finding is unknown. CD117 and SOX-10 expression also occurs in some cutaneous adnexal tumors. This study assesses MYB, CD117 and SOX-10 expression in cutaneous adnexal tumors. METHODS: Retrospective analysis of 184 benign adnexal tumors (140 eccrine/apocrine, 40 follicular and 10 sebaceous), and 30 malignant adnexal tumors was performed with MYB, SOX-10 and CD117 immunostaining. RESULTS: In the benign adnexal tumors, 16% (23/140) significantly expressed MYB. MYB expression was limited to cylindromas and to a lesser extent, spiradenomas in the benign cohort. Elevated MYB expression was detected in mucinous carcinoma, endocrine mucin-producing sweat gland carcinoma and 1 and 4 cases of extramammary Paget's disease (EMPD) in the malignant cohort. CD117 and SOX-10 had similar overall positivity rates in benign apocrine and eccrine tumors (45% and 68% respectively), and were generally negative in other benign and malignant adnexal tumors. CONCLUSION: Expression of MYB appears limited to a small number of cutaneous adnexal tumors, including cylindromas, spiradenomas, ACCs, mucinous carcinoma, endocrine mucin-producing sweat gland carcinoma and some cases of EMPD.

Sox10 and DOG1 Expression in Primary Adnexal Tumors of the Skin.

Primary skin adnexal tumors can be challenging to classify and must be discerned from cutaneous adenocarcinoma metastases from various sites. We evaluated expression of Sox10 and DOG1 in normal cutaneous adnexa and in 194 primary skin adnexal tumors, and compared their performance in discriminating primary skin adnexal tumors from cutaneous metastatic adenocarcinomas with that of p40 and p63. In normal skin adnexa, we noted Sox10 expression in both the secretory and myoepithelial cells in eccrine glands, but only in myoepithelial cells in apocrine glands. DOG1 demonstrated canalicular expression in eccrine glands, and weak expression in myoepithelial cells of apocrine glands, germinative cells of sebaceous glands, and outer root sheath of follicular infundibulum. Sox10 was expressed in 100% of cylindromas and spiradenomas, and in variable frequency in other benign and malignant tumors of sweat glands. DOG1 was positive in most cylindromas (87.5%), in only 10.5% of spiradenomas, and was variably expressed in other benign and malignant tumors of sweat glands. All syringomas (n = 20) were negative for Sox10 and DOG1. One out of the 33 follicular neoplasms was positive for Sox10 and DOG1 (3%). All sebaceous neoplasms were negative for Sox10, and 28.1% of them were positive for DOG1. Sox10 was specific (91.9%) but not sensitive (28.4%) for primary skin origin, and was far less accurate (38.5%) than p63 or p40 (95.5% accuracy). Combining Sox10 with p63 or p40 showed only very minimal gain in accuracy (96%). DOG1 expression in tumors showed low sensitivity and specificity for skin adnexal origin.