Alternate pathways of the early secretory route in yeast

The diversity of functions of eukaryotic cells is preserved by enclosing different enzymatic activities into membrane-bound organelles. Separation of exocytic proteins from those which remain in the endoplasmic reticulum (ER) casts the foundation for correct compartmentalization. The secretory pathway, starting from the ER membrane, operates by the aid of cytosolic coat proteins (COPs). In anterograde transport, polymerization of the COPII coat on the ER membrane is essential for the ER exit of proteins. Polymerization of the COPI coatomer on the cis-Golgi membrane functions for the retrieval of proteins from the Golgi for repeated use in the ER. The COPII coat is formed by essential proteins; Sec13/31p and Sec23/24p have been thought to be indispensable for the ER exit of all exocytic proteins. However, we found that functional Sec13p was not required for the ER exit of yeast endogenous glycoprotein Hsp150 in the yeast Saccharomyces cerevisiae. Hsp150 turned out to be an ATP phosphatase. ATP hydrolysis by a Walker motif located in the C-terminal domain of Hsp150 was an active mediator for the Sec13p and Sec24p independent ER exit. Our results suggest that in yeast cells a fast track transport route operates in parallel with the previously described cisternal maturation route of the Golgi. The fast track is used by Hsp150 with the aid of its C-terminal ATPase activity at the ER-exit. Hsp150 is matured with a half time of less than one minute. The cisternal maturation track is several-fold slower and used by other exocytic proteins studied so far. Operative COPI coat is needed for ER exit by a subset of proteins but not by Hsp150. We located a second active determinant to the Hsp150 polypeptide s N-terminal portion that guided also heterologous fusion proteins out of the ER in COPII coated vesicles under non-functional COPI conditions for several hours. Our data indicate that ER exit is a selective, receptor-mediated event, not a bulk flow. Furthermore, it suggests the existence of another retrieval pathway for essential reusable components, besides the COPI-operated retrotransport route. Additional experiments suggest that activation of the COPI primer, ADP ribosylation factor (ARF), is essential also for Hsp150 transport. Moreover, it seemed that a subset of proteins directly needed activated ARF in the anterograde transport to complete the ER exit. Our results indicate that coat structures and transport routes are more variable than it has been imagined.

Ultrastructural features of the early secretory pathway in Trichoderma reesei

We have systematically analysed the ultra structure of the early secretory pathway in the Trichoderma reesei hyphae in the wild-type QM6a, cellulase overexpressing Rut-C30 strain and a Rut-C30 transformant BV47 overexpressing a recombinant BiP1-VenusYFP fusion protein with an endoplasmic reticulum (ER) retention signal. The hyphae were studied after 24h of growth using transmission electron microscopy, confocal microscopy and quantitative stereological techniques. All three strains exhibited different spatial organisation of the ER at 24h in both a cellulase-inducing medium and a minimal medium containing glycerol as a carbon source (non-cellulase-inducing medium). The wild-type displayed a number of ER subdomains including parallel tubular/cisternal ER, ER whorls, ER-isolation membrane complexes with abundant autophagy vacuoles and dense bodies. Rut-C30 and its transformant BV47 overexpressing the BiP1-VenusYFP fusion protein also contained parallel tubular/cisternal ER, but no ER whorls; also, there were very few autophagy vacuoles and an increasing amount of punctate bodies where particularly the recombinant BiP1-VenusYFPfusion protein was localised. The early presence of distinct strain-specific features such as the dominance of ER whorls in the wild type and tub/cis ER in Rut-C30 suggests that these are inherent traits and not solely a result of cellular response mechanisms by the high secreting mutant to protein overload.

Pathological changes at the target tissue level in Sjögren's syndrome and their effect on the exocrine function of the salivary glands

Sjögren s syndrome (SS) is a common autoimmune disease affecting the lacrimal and salivary glands. SS is characterized by a considerable female predominance and a late age of onset, commonly at the time of adreno- and menopause. The levels of the androgen prohormone dehydroepiandrosterone-sulphate (DHEA-S) in the serum are lower in patients with SS than in age- and sex-matched healthy control subjects. The eventual systemic effects of low androgen levels in SS are not currently well understood. Basement membranes (BM) are specialized layers of extracellular matrix and are composed of laminin (LM) and type IV collagen matrix networks. BMs deliver messages to epithelial cells via cellular LM-receptors including integrins (Int) and Lutheran blood group antigen (Lu). The composition of BMs and distribution of LM-receptors in labial salivary glands (LSGs) of normal healthy controls and patients with SS was assessed. LMs have complex and highly regulated distribution in LSGs. LMs seem to have specific tasks in the dynamic regulation of acinar cell function. LM-111 is important for the normal acinar cell differentiation and its expression is diminished in SS. Also LM-211 and -411 seem to have some acinar specific functional tasks in LSGs. LM-311, -332 and -511 seem to have more general structure maintaining and supporting roles in LSGs and are relatively intact also in SS. Ints α3β1, α6β1, α6β4 and Lu seem to supply structural basis for the firm attachment of epithelial cells to the BM in LSGs. The expression of Ints α1β1 and α2β1 differed clearly from other LM-receptors in that they were found almost exclusively around the acini and intercalated duct cells in salivons suggesting some type of acinar cell compartment-specific or dominant function. Expression of these integrins was lower in SS compared to healthy controls suggesting that the LM-111 and -211-to-Int α1β1 and α2β1 interactions are defective in SS and are crucial to the maintenance of the acini in LSGs. DHEA/DHEA-S concentration in serum and locally in saliva of patients with SS seems to have effects on the salivary glands. These effects were first detected using the androgen-dependent CRISP-3 protein, the production and secretion of which were clearly diminished in SS. This might be due to the impaired function of the intracrine DHEA prohormone metabolizing machinery, which fails to successfully convert DHEA into its active metabolites in LSGs. The progenitor epithelial cells from the intercalated ductal area of LSGs migrate to the acinar compartment and then undergo a phenotype change into secretory acinar cells. This migration and phenotype change seem to be regulated by the LM-111-to-Int α1β1/Int α2β1 interactions. Lack of these interactions could be one factor limiting the normal remodelling process. Androgens are effective stimulators of Int α1β1 and α2β1 expression in physiologic concentrations. Addition of DHEA to the culture medium had effective stimulating effect on the Int α1β1 and α2β1 expression and its effect may be deficient in the LSGs of patients with SS.

Prediction of breakout noise from an elliptical duct of finite length

This paper describes a predictive model for breakout noise from an elliptical duct or shell of finite length. The transmission mechanism is essentially that of ``mode coupling'', whereby higher structural modes in the duct walls get excited because of non-circularity of the wall. Effect of geometry has been taken care of by evaluating Fourier coefficients of the radius of curvature. The noise radiated from the duct walls is represented by that from a finite vibrating length of a semi infinite cylinder in a free field. Emphasis is on understanding the physics of the problem as well as analytical modeling. The analytical model is validated with 3-D FEM. Effects of the ovality, curvature, and axial terminations of the duct have been demonstrated. (C) 2010 Institute of Noise Control Engineering.

Analytical solution of the laminar mean flow wave equation in a lined or unlined two-dimensional rectangular duct

The prediction of the sound attenuation in lined ducts with sheared mean flow has been a topic of research for many years. This involves solving the sheared mean flow wave equation, satisfying the relevant boundary condition. As far as the authors' knowledge goes, this has always been done using numerical techniques. Here, an analytical solution is presented for the wave propagation in two-dimensional rectangular lined ducts with laminar mean flow. The effect of laminar mean flow is studied for both the downstream and the upstream wave propagation. The attenuation values predicted for the laminar mean flow case are compared with those for the case of uniform mean flow. Analytical expressions are derived for the transfer matrices.

Noise reduction with perforated three-duct muffler components

This paper describes the authors’ distributed parameter approach for derivation of closed-form expressions for the four-pole parameters of the perforated three-duct muffler components. In this method, three simultaneous second-order partial differential equations are first reduced to a set of six first-order ordinary differential equations. These equations are then uncoupled by means of a modal matrix. The resulting 6 × 6 matrix is reduced to the 2 × 2 transfer matrix using the relevant boundary conditions. This is combined with transfer matrices of other elements (upstream and downstream of this perforated element) to predict muffler performance like noise reduction, which is also measured. The correlation between experimental and theoretical values of noise reduction is shown to be satisfactory.

On the cut‐on frequencies of a large round duct with azimuthal as well as radial partitions

This letter deals with a three‐dimensional analysis of circular sectors and annular segments resulting from the partitioning of a round (cylindrical) duct for use in an active noise control system. The relevant frequency equations are derived for stationary medium and solved numerically to arrive at the cut‐on frequencies of the first few modes. The resultant table indicates among other things that azimuthal partitioning does not raise the cutoff frequency (the smallest cut‐on frequency) beyond a particular value, and that radial partitioning is counterproductive in that respect.

Analysis of a hybrid noise control system for a duct

This letter proposes the combination of a passive muffler and an active noise control system for the control of very high‐level noise in ducts used with large industrial fans and similar equipment. The analysis of such a hybrid system is presented making use of electroacoustic analogies and the transfer matrix method. It turns out that a passive muffler upstream of the input microphone can indeed lower the acoustic pressure and, hence, the power requirement of the auxiliary source. The parameter that needs to be optimized (or maximized) for this purpose is a certain velocity ratio that can readily be evaluated in a closed form, making it more or less straightforward to synthesize the configuration of an effective passive muffler to go with the active noise control system.

Prediction of Breakout Noise from a Rectangular Duct with Compliant Walls

Breakout noise from HVAC ducts is important at low frequencies, and the coupling between the acoustic waves and the structural waves plays a critical role in the prediction of the transverse transmission loss. This paper describes the analytical calculation of breakout noise by incorporating three-dimensional effects along with the acoustical and structural wave coupling phenomena. The first step in the breakout noise prediction is to calculate the inside duct pressure field and the normal duct wall vibration by using the solution of the governing differential equations in terms of Green's function. The resultant equations are rearranged in terms of impedance and mobility, which results in a compact matrix formulation. The Green's function selected for the current problem is the cavity Green's function with modification of wave number in the longitudinal direction in order to incorporate the terminal impedance. The second step is to calculate the radiated sound power from the compliant duct walls by means of an ``equivalent unfolded plate'' model. The transverse transmission loss from the duct walls is calculated using the ratio of the incident power due to surface source inside the duct to the acoustic power radiated from the compliant duct walls. Analytical results are validated with the FE-BE numerical models.