2 These authors made an equal contribution. Basement membrane molecules such as laminin are important structural components of the skin 1-4 , but also serve as substrates for sensory neurons of the dorsal root ganglia (DRG) to grow in culture 5 . The main function of sensory neurons innervating the skin is to detect and relay relevant sensory stimuli, in particular mechanical stimuli 6 . It has long been known that sensory neurons with a nociceptive function (detecting potentially harmful stimuli) can have their endings in the epidermis 7-9 whereas mechanoreceptor endings (touch receptors) reside exclusively in the dermal layer [9][10] . Interestingly, the matrix environments of the epidermis and the dermis are very distinctive 11 . We showed that mechanosensitive currents required for touch receptor function depend on the presence of a protein tether which may function to couple mechanosensitive channels to a laminin-containing matrix 12 . The tether protein is not required for the mechanosensitivity of most nociceptive sensory neurons. Here we set out to address the idea that sensory mechanotransduction might be modulated by distinct matrix components made by different types of skin cells in different skin layers. We show that epidermal keratinocytes produce a matrix that is non-permissive for mechanotransduction and identify the factor responsible as laminin-332 (formerly known as laminin-5). Laminin matrices doped with small amounts of laminin-332 have a dramatically altered network structure that is non-permissive for tether attachment.We demonstrate a spatially restricted loss of mechanotransduction in neurite segments connected to laminin-332-containing matrices. Mutations in all three genes coding the trimeric laminin-332 protein complex can cause epidermolysis bullosa, a severe inherited skin blistering disease 1,3 . Human keratinocytes that produce a laminin-332 free matrix have no inhibitory activity on mechanotransduction. We have also discovered an activity of laminin-332 matrix in inhibiting sensory axon bifurcation. Our results reveal novel mechanisms whereby permissive and non-permissive substrates can spatially coordinate mechanotransduction in distinct domains within a single neuron. Results Keratinocyte matrix is suppresses mechanotransductionUsing whole-cell, patch-clamp techniques we directly recorded mechanosensitive currents in cultured sensory neurons [12][13][14][15][16][17][18][19][20] . We first asked whether co-culture of sensory neurons with different cellular components of the skin can modulate the activity of mechanosensitive currents. When sensory neurons are cultured on a laminin substrate, standardly-derived from Engelbreth-Holm-Swarm cells (EHS matrix, henceforth referred to as laminin), more than 90% of the cells exhibit a mechanosensitive current evoked using a small (~740 nm displacement) stimulus to the neurite 12, 14-15 . At least three types of mechanosensitive current can be measured in sensory neurons, classified according to their inactivation time constant τ 1 , rap...
Transmission of the malaria parasite from the mammalian host to the mosquito vector requires the formation of adequately adapted parasite forms and stage-specific organelles. Here we show that formation of the crystalloid-a unique and short-lived organelle of the Plasmodium ookinete and oocyst stage required for sporogonyis dependent on the precisely timed expression of the S-acyltransferase DHHC10. DHHC10, translationally repressed in female Plasmodium berghei gametocytes, is activated translationally during ookinete formation, where the protein is essential for the formation of the crystalloid, the correct targeting of crystalloid-resident protein LAP2, and malaria parasite transmission.T he malaria parasite is capable of infecting both the vertebrate host and mosquito vector. After a mosquito blood meal, sexual precursor cells rapidly differentiate into mature gametes. In the mosquito midgut, the gametes mate to form a zygote that develops further into the motile ookinete. After crossing the midgut epithelium and establishing a sessile oocyst, the ookinete gives rise to thousands of sporozoites capable of infecting a subsequent mammalian host (1).Sharing key organelles like the nucleus, endoplasmic reticulum, Golgi, and mitochondria with other eukaryotes, this parasite has evolved specialized, stage-specific structures that are necessary for developmental progression during parasite transmission. These include, for example, osmiophilic bodies (secretory vesicles) that release protein factors capable of lysing the parasitophorous vacuole and erythrocyte membranes, thus producing free gametes (2) and a gliding motility motor anchored to the inner membrane complex (IMC), allowing the ookinete to migrate across the mosquito midgut epithelium and establish an oocyst (3). Sporozoite formation in the oocyst finally requires the presence of a stage-specific organelle, the crystalloid, a multivesicular structure assembled in the ookinete and putative reservoir of proteins and lipids used during sporogony. Although this enigmatic organelle was discovered more than 40 y ago, its formation and function remain largely unknown (4-9). Six LCCL proteins have been shown to reside within (9) and maintain the stability (8, 9) of these organelles essential for sporogony (10).The morphological changes taking place during zygote-toookinete development and the generation of thousands of sporozoites inside a single oocyst require extensive protein translation and membrane biogenesis to support the formation of organelles and plasma membrane (PM) surrounding each new parasite. Onethird of the proteins identified in the oocyst and oocyst-derived (midgut) sporozoites of the human parasite Plasmodium falciparum are putatively membrane-bound (11). The targeting of such proteins to organelles, and perhaps formation of certain organelles per se, requires appropriate sorting signals, along with transmembrane (TM) domains to keep these factors in place. Posttranslational modifications, such as lipidation, can increase the affinity of a modif...
Difficulties in calculating the motor nerve conduction velocity of the peroneal nerve arise when the extensor digitorum brevis is completely atrophied and does not respond to stimulation. I n these patients, conduction velocity i n the proximal fibers of the peroneal nerve can still be calculated by recording from proximal muscles such as the anterior tibial and peroneus brevis. We established normalvalues by asimple standardized method using 34 subjects.Devi S, Lovelace RE, Duarte N: Proximal peroneal nerve conduction velocity: recording from anterior tibial and peroneus brevis muscles. Ann Neurol 2: [116][117][118][119] 1977 The MethodTo establish norms, conduction velocities of the peroneal nerve were determined by recording at the anterior tibial, peroneus brevis, and extensor digitorum brevis muscles. Members of the staff and their families, without neuromuscular disease, constituted the normal group. All studies were performed using a two-channel electromyograph (TECA Model TE4) with fiberoptic permanent recordings. The stimulus was a square-wave pulse with duration variable between 0.5 and 2 msec delivered with a standard bipolar surface electrode. Recording points over the anterior tibial and peroneus brevis muscles corresponded to an area around the motor point. This point was the position where we obtained the greatest amplitude of the negative phase of the evoked muscle potential (also the steepest deflection). Convenient surface markings, using fixed bony points, were devised for these optimum positions.This procedure enabled us to study both the deep and superficial peroneal nerves. The peroneus longus was rejected in favor of the peroneus brevis because of the difficulty of measuring the distal motor latency accurately. In addition, a longer segment was available for calculating the conduction velocity when the peroneus brevis was used. These studies were performed at constant room temperature (22.2-23.3"C), and due care was taken to prevent leg cooling.For the anterior tibial muscle the recording electrode (a metal plate with a diameter of 32 mm) was placed at the junction of the upper third and lower two-thirds of the line between the tibial tuberosity and the tip of the lateral malleolus. The reference electrode was placed over the medial aspect of the tibia at a position 4 cm distal to the recording electrode (Fig 1). For the peroneus brevis the recording electrode was placed at the junction of the upper two-fifths and lower three-fifths of a line between the head of the fibula and the tip of the lateral malleolus. The reference electrode was placed 4 cm distal on the muscle tendon. The ground was placed o n the subcutaneous surface of the tibia, 3 to 4 cm distal from either reference (Fig 2).Recording from the extensor digitorum brevis was performed according to standard procedures [ l , 51. The peroneal nerve was stimulated from above the head of the fibula, just inside the lateral border of the popliteal space at the level of the midpatella. It was also stimulated below the head of the...
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