Purpose Microsatellite instability (MSI) is a pattern of hypermutation that occurs at genomic microsatellites and is caused by defects in the mismatch repair system. Mismatch repair deficiency that leads to MSI has been well described in several types of human cancer, most frequently in colorectal, endometrial, and gastric adenocarcinomas. MSI is known to be both predictive and prognostic, especially in colorectal cancer; however, current clinical guidelines only recommend MSI testing for colorectal and endometrial cancers. Therefore, less is known about the prevalence and extent of MSI among other types of cancer. Methods Using our recently published MSI-calling software, MANTIS, we analyzed whole-exome data from 11,139 tumor-normal pairs from The Cancer Genome Atlas and Therapeutically Applicable Research to Generate Effective Treatments projects and external data sources across 39 cancer types. Within a subset of these cancer types, we assessed mutation burden, mutational signatures, and somatic variants associated with MSI. Results We identified MSI in 3.8% of all cancers assessed—present in 27 of tumor types—most notably adrenocortical carcinoma (ACC), cervical cancer (CESC), and mesothelioma, in which MSI has not yet been well described. In addition, MSI-high ACC and CESC tumors were observed to have a higher average mutational burden than microsatellite-stable ACC and CESC tumors. Conclusion We provide evidence of as-yet-unappreciated MSI in several types of cancer. These findings support an expanded role for clinical MSI testing across multiple cancer types as patients with MSI-positive tumors are predicted to benefit from novel immunotherapies in clinical trials.

Next‐generation sequencing has aided characterization of genomic variation. While whole‐genome sequencing may capture all possible mutations, whole‐exome sequencing remains cost‐effective and captures most phenotype‐altering mutations. Initial strategies for exome enrichment utilized a hybridization‐based capture approach. Recently, amplicon‐based methods were designed to simplify preparation and utilize smaller DNA inputs. We evaluated two hybridization capture‐based and two amplicon‐based whole‐exome sequencing approaches, utilizing both Illumina and Ion Torrent sequencers, comparing on‐target alignment, uniformity, and variant calling. While the amplicon methods had higher on‐target rates, the hybridization capture‐based approaches demonstrated better uniformity. All methods identified many of the same single‐nucleotide variants, but each amplicon‐based method missed variants detected by the other three methods and reported additional variants discordant with all three other technologies. Many of these potential false positives or negatives appear to result from limited coverage, low variant frequency, vicinity to read starts/ends, or the need for platform‐specific variant calling algorithms. All methods demonstrated effective copy‐number variant calling when evaluated against a single‐nucleotide polymorphism array. This study illustrates some differences between whole‐exome sequencing approaches, highlights the need for selecting appropriate variant calling based on capture method, and will aid laboratories in selecting their preferred approach.

Unique among the phospholipase C isozymes, the recently identified phospholipase C-⑀ (PLC- Comparative sequence analysis of mammalian phospholipase C isozymes revealed a unique ϳ65 amino acid insert within the catalytic core of PLC-⑀ not present in PLC-␤, ␥, ␦, or . A PLC-⑀ construct lacking this region was no longer activated by Rho or G␣ 12/13 but retained regulation by G␤␥ and H-Ras. GTP-dependent interaction of Rho with PLC-⑀ was illustrated in pull-down experiments with GST-Rho, and this interaction was retained in the PLC-⑀ construct lacking the unique insert within the catalytic core. These results are consistent with the conclusion that Rho family GTPases directly interact with PLC-⑀ by a mechanism independent of the CDC25 or RA domains. A unique insert within the catalytic core of PLC-⑀ imparts responsiveness to Rho, which may signal downstream of G␣ 12/13 in the regulation of PLC-⑀, because activation by both Rho and G␣ 12/13 is lost in the absence of this sequence.⑀

The conceptual segregation of G protein-stimulated cell signaling responses into those mediated by heterotrimeric G proteins versus those promoted by small GTPases of the Ras superfamily is no longer vogue. PLC-epsilon, an isozyme of the phospholipase C (PLC) family, has been identified recently and dramatically extends our understanding of the crosstalk that occurs between heterotrimeric and small monomeric GTPases. Like the widely studied PLC-beta isozymes, PLC-epsilon is activated by Gbetagamma released upon activation of heterotrimeric G proteins. However, PLC-epsilon markedly differs from the PLC-beta isozymes in its capacity for activation by Galpha(12/13) - but not Galpha(q) -coupled receptors. PLC-epsilon contains two Ras-associating domains located near the C terminus, and H-Ras regulates PLC-epsilon as a downstream effector. Rho also activates PLC-epsilon, but in a mechanism independent of the C-terminal Ras-associating domains. Therefore, Ca(2+) mobilization and activation of protein kinase C are signaling responses associated with activation of both H-Ras and Rho. A guanine nucleotide exchange domain conserved in the N terminus of PLC-epsilon potentially confers a capacity for activators of this isozyme to cast signals into additional signaling pathways mediated by GTPases of the Ras superfamily. Thus, PLC-epsilon is a multifunctional nexus protein that senses and mediates crosstalk between heterotrimeric and small GTPase signaling pathways.

PLC-⑀ was identified recently as a phosphoinositidehydrolyzing phospholipase C (PLC) containing catalytic domains (X, Y, and C2) common to all PLC isozymes as well as unique CDC25-and Ras-associating domains. Novel regulation of this PLC isozyme by the Ras oncoprotein and ␣-subunits (G␣ 12 ) of heterotrimeric G proteins was illustrated. Sequence analyses of PLC-⑀ revealed previously unrecognized PH and EF-hand domains in the amino terminus. The known interaction of G␤␥ subunits with the PH domains of other proteins led us to examine the capacity of G␤␥ to activate PLC-⑀.