No abstract

The evolutionary history of serine proteases can be accounted for by highly conserved amino acids that form crucial structural and chemical elements of the catalytic apparatus. These residues display nonrandom dichotomies in either amino acid choice or serine codon usage and serve as discrete markers for tracking changes in the active site environment and supporting structures. These markers categorize serine proteases of the chymotrypsin-like, subtilisinlike and a/b-hydrolase fold clans according to phylogenetic lineages, and indicate the relative ages and order of appearance of those lineages. A common theme among these three unrelated clans of serine proteases is the development or maintenance of a catalytic tetrad, the fourth member of which is a Ser or Cys whose side chain helps stabilize other residues of the standard catalytic triad. A genetic mechanism for mutation of conserved markers, domain duplication followed by gene splitting, is suggested by analysis of evolutionary markers from newly sequenced genes with multiple protease domains.

A fundamental property of leukemic stem cells is clonal dominance of the bone marrow microenvironment. Truncation mutations of CSF3R, which encodes the G-CSF receptor (G-CSFR), are implicated in leukemic progression in patients with severe congenital neutropenia. Here we show that expression of a truncated mutant Csf3r in mice confers a strong clonal advantage at the HSC level that is dependent upon exogenous G-CSF. G-CSF-induced proliferation, phosphorylation of Stat5, and transcription of Stat5 target genes were increased in HSCs isolated from mice expressing the mutant Csf3r. Conversely, the proliferative advantage conferred by the mutant Csf3r was abrogated in myeloid progenitors lacking both Stat5A and Stat5B, and HSC function was reduced in mice expressing a truncated mutant Csf3r engineered to have impaired Stat5 activation. These data indicate that in mice, inappropriate Stat5 activation plays a key role in establishing clonal dominance by stem cells expressing mutant Csf3r.

Serine proteases of the chymotrypsin family have maintained a common fold over an evolutionary span of more than one billion years. Notwithstanding modest changes in sequence, this class of enzymes has developed a wide variety of substrate specificities and important biological functions. Remarkably, the C-terminal portion of the sequence in the protease domain accounts fully for this functional diversity. This portion is often encoded by a single exon and contains most of the residues forming the contact surface in the active site for the P1-P3 residues of the substrate, as well as domains responsible for the modulation of catalytic activity. The evolution of serine proteases was therefore driven by optimization of contacts made with the unprimed subsites of the substrate and targeted a relatively short portion of the sequence toward the C-terminal end. The dominant role of the C-terminal sequence should facilitate the identification of function in newly discovered genes belonging to this class of enzymes.Serine proteases of the chymotrypsin family feature a gamut of important physiological functions, ranging from digestive and degradative processes to blood clotting, cellular and humoral immunity, fibrinolysis, fertilization, and embryonic development (1). How this variety of functions emerged during evolution from a highly conserved three-dimensional fold (2) is incompletely understood. Enzymes involved in digestive and degradative processes, such as trypsin and chymotrypsin, are found from bacteria to human and are composed of a protease domain containing all epitopes for ligand recognition (3, 4). On the other hand, enzymes involved in more specialized functions such as blood coagulation, humoral immunity, and fibrinolysis are present almost exclusively in vertebrates and carry additional modules that confer more stringent specificity and localize the proteolytic function in space (5-7). A large repertoire of functions can be created by linking the protease domain to specialized functional modules, and most likely this strategy enabled the evolution of highly selective and specialized enzymes from primitive digestive proteases (8).The mosaic organization of the primary structure of many serine proteases has fostered the notion that the protease domain alone would not bear reliable signatures of function. Previous sequence analyses and functional characterizations of serine proteases have relied heavily on the architecture of non-protease domains (6,8,9). Evolutionary trees for the hepatocyte growth factor (HGF), 1 HGF activator, kallikreins, mast cell protease, and complement lineages have been constructed in this manner (7, 10 -13). However, Doolittle and Feng (5) were able to construct a reasonable evolutionary tree for clotting and fibrinolytic proteases from analysis of the protease domain only. This domain is the only structural component present in all members of the serine protease family. We therefore posed the question as to whether this domain would suffice to gain a predictive understanding of th...

Prior studies suggest increased CMV infection following haploidentical donor transplantation with post-transplant cyclophosphamide (HaploCy). The role of allograft source and PTCy in CMV infection and disease is unclear. We analyzed the effect of graft source and PTCy on incidence of CMV infection as well as transplant outcomes as it relates to CMV serostatus and occurrence of CMV infection by d180. We examined patients reported to CIBMTR between 2012-2017 who had received HaploCy (n = 757), Sib with PTCy (SibCy, n=403), or Sib with calcineurin inhibitor-based prophylaxis (SibCNI, n=1605) for AML/ALL/MDS. Cumulative incidences of CMV infection by d180 were 42% (99% CI, 37-46), 37% (31 - 43), and 23% (20 - 26), respectively [p<0.001]. CMV end-organ disease was statistically comparable. CMV infection risk was highest for CMV-Seropositive recipients (R+), but significantly higher in PTCy recipients regardless of donor [HaploCy (n=545): HR 50.3 (14.4 - 175.2); SibCy (n=279): HR 47.7 (13.3 - 171.4); SibCNI (n=1065): HR 24.4 (7.2 - 83.1); p<0.001]. D+/R- patients also had increased risk for CMV infection. Among seropositive recipients or those developing CMV infection, HaploCy had worse OS and NRM. Relapse was unaffected by CMV infection or serostatus. PTCy was associated with lower chronic GVHD overall, but CMV infection in PTCy recipients was associated with higher cGVHD (p=0.006). PTCy, regardless of donor, is associated with higher incidence of CMV infection, augmenting the risk of seropositivity. Additionally CMV infection may negate the cGVHD protection of PTCy. This study supports aggressive prevention strategies in all patients receiving PTCy.