Ttc39c is upregulated during skeletal muscle atrophy and modulates ERK1/2 MAP kinase and hedgehog signaling.

Tetratricopeptide repeat domain containing 39c (Ttc39c) is expressed in skeletal muscle and is transcriptionally activated in response to neurogenic atrophy in mice. Expression analysis using quantitative polymerase chain reaction and Western blots revealed that Ttc39c is expressed in both proliferating and differentiated muscle cells, peaking during early differentiation and then decreasing as cells progress further through the differentiation process. To further analyze the transcriptional regulation of Ttc39c, promoter fragments of the gene were cloned and fused with the secreted alkaline phosphatase reporter gene. The Ttc39c reporter plasmids were then transfected into cultured mouse muscle cells and found to have transcriptional activity. Furthermore, overexpression of MyoD and myogenin resulted in significant transcriptional repression of the Ttc39c reporter genes. To determine subcellular localization, an expression plasmid with the Ttc39c complementary DNA fused with green fluorescent protein was transfected into muscle cells and analyzed by confocal fluorescent microscopy showing that Tct39c localizes exclusively to the cytoplasm of cultured cells. To assess potential function in muscle, Ttc39c was overexpressed leading to vitiated muscle cell differentiation, impaired ERK1/2 MAP Kinase and Hedgehog signaling, and increased expression of IFT144 and IFT43, which are part of the IFT-A complex involved in retrograde transport in primary cilia. Interestingly, Ttc39c knockdown also resulted in inhibition of muscle cell differentiation and impaired Hedgehog and MAP Kinase signaling but did not affect IFT144 or IFT433 expression. The results of this study demonstrate that muscle cell differentiation is sensitive to abnormal Ttc39c expression and that normal Ttc39c expression appears to be necessary for proper MAP Kinase and Hedgehog signal transduction in developing muscle cells.

Performance of MYC, BCL2, and BCL6 break-apart FISH in small biopsies with large B-cell lymphoma: a retrospective Cytopathology Hematopathology Interinstitutional Consortium study.

Introduction: Fluorescence in situ hybridization (FISH) is an essential ancillary study used to identify clinically aggressive subsets of large B-cell lymphomas that have MYC, BCL2, or BCL6 rearrangements. Small-volume biopsies such as fine needle aspiration biopsy (FNAB) and core needle biopsy (CNB) are increasingly used to diagnose lymphoma and obtain material for ancillary studies such as FISH. However, the performance of FISH in small biopsies has not been thoroughly evaluated or compared to surgical biopsies. Methods: We describe the results of MYC, BCL2, and BCL6 FISH in a series of 222 biopsy specimens, including FNAB with cell blocks, CNBs, and surgical excisional or incisional biopsies from 208 unique patients aggregated from 6 academic medical centers. A subset of patients had FNAB followed by a surgical biopsy (either CNB or excisional biopsy) obtained from the same or contiguous anatomic site as part of the same clinical workup; FISH results were compared for these paired specimens. Results: FISH had a low hybridization failure rate of around 1% across all specimen types. FISH identified concurrent MYC and BCL2 rearrangements in 20 of 197 (10%) specimens and concurrent MYC and BCL6 rearrangements in 3 of 182 (1.6%) specimens. The paired FNAB and surgical biopsy specimens did not show any discrepancies for MYC or BCL2 FISH; of the 17 patients with 34 paired cytology and surgical specimens, only 2 of the 49 FISH probes compared (4% of all comparisons) showed any discrepancy and both were at the BCL6 locus. One discrepancy was due to necrosis of the CNB specimen causing a false negative BCL6 FISH result when compared to the FNAB cell block that demonstrated a BCL6 rearrangement. Discussion: FISH showed a similar hybridization failure rate in all biopsy types. Ultimately, MYC, BCL2, or BCL6 FISH showed 96% concordance when compared across paired cytology and surgical specimens, suggesting FNAB with cell block is equivalent to other biopsy alternatives for evaluation of DLBCL or HGBCL FISH testing.