The conversion of light to chemical energy by the process of photosynthesis is localized to the thylakoid membrane network in plant chloroplasts. Although several pathways have been described that target proteins into and across the thylakoids, little is known about the origin of this membrane system or how the lipid backbone of the thylakoids is transported and fused with the target membrane. Thylakoid biogenesis and maintenance seem to involve the flow of membrane elements via vesicular transport. Here we show by mutational analysis that deletion of a single gene called VIPP1 (vesicle-inducing protein in plastids 1) is deleterious to thylakoid membrane formation. Although VIPP1 is a hydrophilic protein it is found in both the inner envelope and the thylakoid membranes. In VIPP1 deletion mutants vesicle formation is abolished. We propose that VIPP1 is essential for the maintenance of thylakoids by a transport pathway not previously recognized.

Knudson's two-hit hypothesis postulates that genetic alterations in both alleles are required for the inactivation of tumor-suppressor genes. Genetic alterations include small or large deletions and mutations. Over the past years, it has become clear that epigenetic alterations such as DNA methylation are additional mechanisms for gene silencing. Restriction Landmark Genomic Scanning (RLGS) is a two-dimensional gel electrophoresis that assesses the methylation status of thousands of CpG islands. RLGS has been applied successfully to scan cancer genomes for aberrant DNA methylation patterns. So far, the majority of this work was done using NotI as the restriction landmark site. Here, we describe the development of RLGS using AscI as the restriction landmark site for genome-wide scans of cancer genomes. The availability of AscI as a restriction landmark for RLGS allows for scanning almost twice as many CpG islands in the human genome compared with using NotI only. We describe the development of an AscI–EcoRV boundary library that supports the cloning of novel methylated genes. Feasibility of this system is shown in three tumor types, medulloblastomas, lung cancers, and head and neck cancers. We report the cloning of 178 AscI RLGS fragments via two methods by use of this library.[Supplemental material is available online at http://www.genome.org.]

All plastomes of land plants and alga analyzed to date encode homologues of the core subunits of an eubacterialike RNA polymerase. However, the plastomes lack genes for sigma factors which are imperative for proper promoter recognition by this type of RNA polymerase and hence must be encoded by nuclear genes. By searching plant "expressed sequence tags" (EST5) for similarities to eubacterial sigma factors an Arabidopsis EST (155H23T7) was identified which was used for the isolation of a homologous full-length cDNA from the C4 monocot, Sorghum bicolor. The cDNA sequence encodes a protein exhibiting the typical sequence motives of cs70-type sigma factors and the corresponding gene was therefore designated Sigi. The SIG1 protein shows the greatest similarity to recently identified plastidial sigma factors of A. thaliana, 0. sativa and S. alba. Like these proteins the Sorghum SIG1 protein contains an aminoterminal extension reminiscent of plastidial transit sequences indicating that the SIG1 protein functions as a plastidial sigma factor. Accordingly, Sigi transcripts are found preferentially in leaves and accumulate in a light-dependent manner. Sigi transcript levels are similar in the mesophyll and bundlesheath cells suggesting that at least this sigma factor is not involved in the differential expression of plastid genes in the two cell types. Southern analyses indicate that the Sigl sequence is present as single copy gene in the Sorghum genome.

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