Head-to-tail macrocyclization of cysteine-free peptides using an o-aminoanilide linker

A head-to-tail macrocyclization protocol for the preparation of cysteine-free cyclic peptides was investigated. The o-aminoanilide linker constructed in the peptide sequence by a standard Fmoc-based peptide synthesis procedure was subjected to nitrite-mediated activation under acidic conditions toward N-acyl benzotriazole as the active ester species. The subsequent cyclization smoothly proceeded by neutralization in the presence of additives such as 1-hydroxybenzotriazole (HOBt) and 1-hydroxy-7-azabenzotriazole (HOAt) to afford the expected cyclic pentapeptide, a CXCR4 antagonist. The cyclization efficiencies were dependent on the precursor open-chain sequence. The application of this step-wise activation-cyclization protocol to microflow reaction systems is also described.     

CPAP promotes timely cilium disassembly to maintain neural progenitor pool

A mutation in the centrosomal‐P4.1‐associated protein (CPAP) causes Seckel syndrome with microcephaly, which is suggested to arise from a decline in neural progenitor cells (NPCs) during development. However, mechanisms of NPCs maintenance remain unclear. Here, we report an unexpected role for the cilium in NPCs maintenance and identify CPAP as a negative regulator of ciliary length independent of its role in centrosome biogenesis. At the onset of cilium disassembly, CPAP provides a scaffold for the cilium disassembly complex (CDC), which includes Nde1, Aurora A, and OFD1, recruited to the ciliary base for timely cilium disassembly. In contrast, mutated CPAP fails to localize at the ciliary base associated with inefficient CDC recruitment, long cilia, retarded cilium disassembly, and delayed cell cycle re‐entry leading to premature differentiation of patient iPS‐derived NPCs. Aberrant CDC function also promotes premature differentiation of NPCs in Seckel iPS‐derived organoids. Thus, our results suggest a role for cilia in microcephaly and its involvement during neurogenesis and brain size control.     

Delayed fluorescence observed in the nanosecond time region at 77 K originates directly from the photosystem II reaction center

The excited-state dynamics of delayed fluorescence in photosystem (PS) II at 77 K were studied by time-resolved fluorescence spectroscopy and decay analysis on three samples with different antenna sizes: PS II particles and the PS II reaction center from spinach, and the PS II core complexes from Synechocystis sp. PCC 6803. Delayed fluorescence in the nanosecond time region originated from the 683-nm component in all three samples, even though a slight variation in lifetimes was detected from 15 to 25 ns. The relative amplitude of the delayed fluorescence was higher when the antenna size was smaller. Energy transfer from the 683-nm pigment responsible for delayed fluorescence to antenna pigment(s) at a lower energy level was not observed in any of the samples examined. This indicated that the excited state generated by charge recombination was not shared with antenna pigments under the low-temperature condition, and that delayed fluorescence originates directly from the PS II reaction center, either from chlorophyll a(D1) or P680. Supplemental data on delayed fluorescence from spinach PS I complexes are included.     

Immunological characterization of human vitronectin and its binding to glycosaminoglycans

The cell-adhesive glycoprotein vitronectin in human plasma was characterized with a monospecific anti-vitronectin antibody. Vitronectin, a mixture of monomeric 75 and 65 kDa polypeptides, was found to have different ratios of amounts of 75 and 65 kDa polypeptides in immunoblots of sera from various healthy human donors. Two states of vitronectin were previously reported; the open state binds to heparin, but the cryptic state does not (Hayashi et al. (1985) J. Biochem. 98, 1135-1138). The anti-vitronectin antibody was suggested to react more strongly with the open state of vitronectin than with the cryptic state. To quantitate all vitronectin regardless of its state, an enzyme-linked immunosorbent assay of vitronectin was developed based on prior boiling of vitronectin-containing samples in 2% (w/v) sodium dodecyl sulfate and 40 mM dithiothreitol to destroy conformational differences. About 12-20% of the vitronectin molecules in plasma were found to bind to heparin-Sepharose under physiological conditions. Vitronectin in plasma bound 30-fold more efficiently to heparin immobilized by amino groups than by carboxyl groups. Its affinity for heparin was higher than for chondroitin sulfate A or C, or dermatan sulfate. Vitronectin was also found to contain covalently-linked small polypeptides of 15 and 13 kDa. These light chains seemed to be disulfide-bonded to the 65 kDa polypeptide, and might be endogenously derived from nicks in the carboxy-terminal portion of the 75 kDa polypeptide in plasma.     

New information on age and growth in length of Micromesistius australis,Norman 1937 (Pisces,Gadidae), in the South-West Atlantic

Summary Sagittal otoliths were used for age determination in 1121 specimens of Micromesistius australis, caught in waters of the South-West Atlantic during the cruises made by the ships Walther Herwig and Shinkai Maru in agreement with the Argentine Republic, during the years 1978 and 1978/1979, respectively. The interpretation of growth rings required the otoliths to undergo a paraffin wax embedding and cutting technique. The relations otolith length-fish length and fish length-half otolith width were linear. Thus, back calculations, when necessary, were made by a simple extrapolation. Numbers of males and females were nearly equal, resulting in a sex ratio of 1:1.09. Length frequencies and age distributions were calculated for each sex separately. A significant difference in growth between sexes was found, curves for each were derived. A linear, regression was made and the Von Bertalanffy equation estimated as: Lt=56.90 [1-e^{-0.1983 (t+1.84)}], in males, and Lt=60.08 [1-e^{-0.1769 (t+2.07)}] in females, with Lt in cm and t in years. For estimation of these equation parameters age groups 2–16 years in males and 2–22 years in females were taken into account. All theoretical Lt values and almost all observed lengths for females were greater than those of males in corresponding age groups. Previously published observations about M. australis on this subject, are not in agreement or only in partial accord with present results. These are discussed. Original photographs of otolith cross sections illustrating age determination methods are presented for the first time in age and growth studies of this species.     

Tumoricidal activity of Kupffer cells augmented by anticancer drugs.

The effect of Mitomycin-C (MMC) and Adriamycin (ADM) on the antitumor-associated function of Kupffer cells was examined. MMC and ADM enhanced the production of superoxide by Kupffer cells in cultures at low concentrations likely to occur in clinical use. The expression of interleukin-2 receptor, Ia antigen and asialoGM1 antigen, measured by flowcytometry, was increased by contact with MMC. Growth inhibition of AH130, rat ascites hepatoma, and P815, murine mastocytoma, by Kupffer cells treated with anticancer drugs was greater than that by Kupffer cells alone or anticancer agent alone. These results show that MMC and ADM activate Kupffer cells, leading to synergistic antitumor activity. The results suggest that some anticancer agents act through immunological mechanisms as well as through direct antineoplastic activity.     

Enhancement of rat hepatic macrophages by treatment with interleukin-2 and streptococcal preparation OK432, with reference to antitumor activity,soluble factor production and Ia expression

Summary The effect of biological response modifiers, such as interleukin-2 (IL-2) and streptococcal preparation OK432, on the functions of hepatic macrophages was investigated. The macrophages, even with no exogenous stimulation, produced superoxide anion (O ₂ ^- ) and tumor necrosis factor (TNF), displayed cytotoxicity against K562 cells and cytostasis against P815 cells and expressed immune-region-associated antigen (Ia). IL-2 administered in vitro or in vivo enhanced O ₂ ^- production by hepatic macrophages and the intravenous injection of OK432 also enhanced O ₂ ^- production. Furthermore, IL-2 added to the culture medium of hepatic macrophages isolated from OK432-injected rats augmented O ₂ ^- production even more. The TNF production and Ia expression of the macrophages were also increased by the intravenous injection of OK432. As with O ₂ ^- production, the cytotoxicity of the cells was enhanced by OK432 injection or by IL-2 added to the culture medium and the combination of OK432 and IL-2 augmented their cytotoxicity even more. Thus, the present study suggested that IL-2 and OK432 induce the augmentation of the antitumor activity of hepatic macrophages, partly as a result of the increase in production of O ₂ ^- and TNF and Ia expression.